Review Article

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Ann Phlebology 2022; 20(2): 68-77

Published online December 31, 2022

https://doi.org/10.37923/phle.2022.20.2.68

© Annals of phlebology

New Treatment of Varicose Veins through Muscle Regeneration of Lower Leg Muscles (Especially Calf Muscle) Without Removal of Varicose Veins

Ki Ji Lee, M.D., Ph.D.

Department of Pediatrics, Woorisoa Hospital, Seoul, Korea

Correspondence to : Ki Ji Lee, 15 Saemal-ro, Guro-gu, Seoul 47848, Korea, Deparment of Pediatrics, Woorisoa Seoul Hospital
Tel: 02-858-0100, Fax: 02-856-0032
E-mail: leekiji@naver.com

The calf muscle pump is the motive force enhancing venous blood return from the lower extremity to the heart. It causes displacement of venous blood in both vertical and horizontal directions, generates ambulatory pressure gradient between the thigh and lower leg veins, and bidirectional streaming within calf perforators. Ambulatory pressure gradient triggers venous reflux in incompetent veins, inducing ambulatory venous hypertension in the lower leg and foot. Bidirectional flow in calf perforators enables quick pressure equalization between deep and superficial veins of the lower leg; the outward (into the superficial veins) oriented component of the bidirectional flow taking place during calf muscle contraction is not a pathological reflux but a physiological centripetal flow streaming via the great saphenous vein into the femoral vein. Calf perforators are communicating channels between both systems, making them conjoined vessels; they are not involved in generating pathological hemodynamic situations and do not cause ambulatory venous hypertension. Pressure gradient arising during calf pump activity between the femoral vein and the saphenous remnant after abolishing saphenous reflux triggers biophysical and biochemical events, which might induce recurrence. Thus, abolishing saphenous reflux removes the hemodynamic disturbance but simultaneously generates a precondition for reflux recurrence and the return of the previous pathological situation; this chain of events has been termed the hemodynamic paradox. But this review showed that varicose veins could be improved quickly through lower leg muscles (especially calf muscle) regeneration by increasing mitochondrial cellular energy (adenosine triphosphate) of leg muscles without removing varicose veins.

Keywords Calf muscle pump, Venous hemodynamics, Muscle regeneration, Muscle stem cells (satellite cells), Mitochondrial energy (adenosine triphosphate)

Vis a tergo, the remainder of the cardiac energy propelling venous blood from the lower extremities against the gravitation toward the heart is not powerful enough to move the venous blood toward the heart effectively; the pressure difference between the venules and the right atrium is approximately 20 mmHg. In addition to this motive force, the calf muscle pump (CMP) constitutes an effective mechanism that promotes the drainage of venous blood from the lower extremity, produces a marked increase of systolic pressure, decreases diastolic pressure in deep and superficial veins of the lower leg, and entails a distinct physiological decrease of ambulatory venous pressure (AVP) in the lower leg veins. The requirements for efficiency and effectiveness of the CMP are patency and perfect competence of deep lower leg veins, no impedi-ments in the venous outflow tract, and undisturbed contractility of the calf musculature. The venous reflux compromises the physiological effectivity of the CMP since the venous blood volume expelled by the CMP is more or less replaced by the refluxing volume. The contribution of the foot pump to the venous return from the lower extremity remains clear. This review article intends to evaluate the physiological significance of the CMP and the foot pump, assess the systolic and diastolic pressure behavior in deep and superficial veins of the lower leg during calf pump activity, and evaluate the flow behavior in calf perforators resulting from systolic and diastolic pressure differences.

The calf musculature and the deep lower leg veins form the CMP. The calf musculature consists of

- 1. The posterior compartment inducing plantar flexion of the ankle joint and the toes, as well as supination of the foot.

- 2. The anterior compartment inducing dorsal flexion of the ankle joint and the toes.

- 3. The lateral compartment inducing plantar flexion of the ankle joint and pronation of the foot.

Deep lower leg veins consist of three paired conductive veins: the posterior tibial vein (PTV), anterior tibial vein, peroneal vein, and the muscular veins contained in the sural musculature (most important: gastrocnemius and soleus veins). The CMP is called “peripheral heart”: contractions of the sural muscles eject the venous blood centripetally toward the heart and promote venous blood return from the lower extremity against the gravitation force substantially. During calf muscle contraction, the pressure rises conside-rably in deep lower leg veins (on average of 75 mmHg) and a little less in the great saphenous vein (GSV). The systolic pressure difference between the PTV and GSV amounts to approximately 40 mmHg in healthy individuals and merely 14 mmHg in patients with varicose veins (1-3); the unequal flow rate capacity of the calf perforators in healthy individuals and patients with varicose vein causes this pressure gradient difference. Thus, during calf muscle contractions, the venous blood is ejected mainly into the popliteal vein, but a smaller part escapes through calf perforators into the GSV and streams further in the centripetal direction toward the heart. During calf muscle relaxation, the diastolic pressure decreases in deep lower leg veins and GSV; the pressure gradient reverses, and the diastolic pressure in the GSV is approximately 13 mmHg higher than in the PTV (2,3). The systolic and diastolic pressure changes are produced in deep veins. Pressure transmissions between the PTV and GSV occur through the connecting channels, through calf perforators, documented by simultaneous pressure recordings. The pressure trans-missions could not occur without bidirectional streaming through calf perforators, i.e., without systolic outward and diastolic inward flows. The outward flow produces a steep systolic increase in pressure in the GSV through calf perforators. This flow is centripetal, i.e., physiological; it continues via GSV toward to heart and can be assessed in the GSV by duplex ultrasonography (DUS). In healthy individuals with competent calf perforators, there was a steep increase in systolic pressure in the GSV; however, a similar steep increase in the PTV could not occur. Similar conditions exist in the heart across the competent and incompetent mitral valve. A competent mitral valve precludes a systolic increase in pressure in the left atrium. Regurgitation through an incompetent mitral valve induces increased pressure in the left atrium. Thus, with some benevolence, CMP can be compared with the left ventricle and the saphenous system with the left atrium. The pressure in the popliteal and femoral vein oscillates during the CMP activity but does not decrease below the hydrostatic pressure value during the diastolic phase (1,2,4). The pressure difference arising in this way between the popliteal/femoral veins and deep lower leg veins has been called ambulatory pressure gradient; its value is 37.4±6.4 mmHg (5). Ambulatory pressure gradient triggers reflux in incompetent veins connecting both poles of the pressure gradient. The performance of the CMP depends primarily on the patency and impeccable competence of the deep conductive veins of the lower leg (posterior and anterior tibial veins and peroneal veins). The incompetence of these veins disturbs the performance of the CMP. Further factors impairing CMP performance are obstacles in the outflow venous tracts and reduced calf muscle strength due to muscle weakness, atrophy, or palsy. Plethysmographic findings enable quantification of the CMP performance. Ejection volume produced by the CMP is an important parameter indicating its effectivity; it is determined by air-plethysmography; the normal values range from 60 to 90 ml (5). Ejection fraction is another parameter used for evaluating CMP performance; it is expressed in percentage and is calculated as the volume of blood ejected with one calf muscle contraction divided by the functional venous volume. Since this parameter depends on the value of the functional venous volume, it is not suitable for evaluating results after therapeutic procedures that reduce the venous volume contained in varicose veins. The ejection fraction, i.e., the quotient EV/VV, improves and simulates improved calf muscle performance, but the pumping effectivity remains unaffected. Other valuable plethysmographic para-meters registered after a series of calf muscle contractions and relaxations, such as refill time t-50 and t-90, refill volume gained by strain gauge plethysmography, and residual volume gained by air plethysmography, are influenced by the CMP performance and venous reflux. Saphenous reflux simulates CMP malfunction because the refluxing volume devaluates the ejection volume. Therefore, the last mentioned parameters can be considered for evaluating the CMP performance only after eliminating saphenous reflux. As a rule, in patients with primary varicose veins, even the severest hemodynamic disorders in clinical, etiological, anatomical, and pathological classes C4-C6 are caused by saphenous reflux, whereas the performance of the CMP remains unaffected. The elimina-tion of saphenous reflux restores physiological AVP conditions and normal plethysmographic values, which would be impossible if CMP performance were impaired (6-9).

The plantar venous plexus forms a substantial part of the venous reservoir of the foot pump. According to Binns and Pho (10), the plantar venous plexus consists, in addition to subtle veins, mainly of the large lateral plantar vein that is partly double-barreled, of a little smaller medial plantar vein and the deep plantar venous arch. The plantar veins are convoluted and fitted with valves directing the flow into the PTV. From the diameter and length of these veins, it can be computed that the plantar venous plexus contains approximately 6 ml of blood. In another study, White et al. (11) reported an average of 2.7 large veins in the plantar venous plexus with a diameter of 4.0±1.0 mm. The entire length of these veins along the forefoot was approximately 30 cm, meaning they contained approximately 5 ml of blood. So, the total venous volume of the sole, including the blood volume in the foot muscles, might hardly reach 10 ml. The contribution of the foot pump to the venous return is still disputable. Plantar veins empty due to compression exerted by weight bearing. Due to the foot’s anatomical configuration, only the lateral part is involved in the pumping process. When the normal ejection fraction of 60% is considered, the estimated ejection volume of the foot pump would be approximately 3∼4 ml; the large part of this very small volume is ejected into deep lower leg veins, and a little part might be ejected into the GSV. Broderick et al. (12) researched venous emptying from the foot using several provoking maneuvers: weight bearing, toe curl, electrical stimulation of the tibial nerve at the medial malleolus, and passive compression. Flow measurements in the particular lower leg veins (posterior tibial, anterior tibial, and peroneal veins) and popliteal veins were performed using DUS; the flow in the GSV was not measured. Flow measurements in the deep lower leg veins were performed at the level between the lower and middle part of the lower leg. Values of the ejection volume were obtained by multiplying the cross-sectional area by the mean velocity and total flow duration. Electrical stimulation of the foot muscles ejected just 1 ml of venous blood into the lower leg veins altogether, indicating that the contraction of the metatarsal muscles exerts just a little effect. Intermittent pneumatic foot compression produced a venous outflow of 2.18 ml, which is likewise negligible. Weight-bearing maneuver expelled 6.08 ml and toe curls 5.03 ml of venous blood into deep lower leg veins. Oddly enough, weight-bearing maneuvers and toe curls induced in the popliteal vein a remarkable centripetal flow of 33.34 ml and 31.04 ml, respectively, showing that weight-bearing maneuvers and toe curls activated the foot pump and, more effectively also, the CMP. It is quite difficult to selectively provoke the foot pump activity without evoking in the weigh-bearing limb involuntary contractions of the calf musculature that occur to ensure the balance of the body. In addition, toe curls activate the CMP (musculus flexor hallucis longus and musculus flexor digitorum longus). Since the flow measurements in the lower leg veins were performed at the level between the lower and middle third of the lower leg, it can be deduced that the upper section of the calf musculature where the substantial mass of calf musculature is concentrated might be the most important part of the CMP. The findings documented that intermittent pneumatic compression of the sole and active contractions of the foot muscles could not expel a significant volume of venous blood from the foot. The total volume ejected by these two maneuvers into deep lower leg veins amounted to 3.18 ml (1 ml due to electrical stimulation of the tibial nerve, 2.18 ml due to intermittent pneumatic compression), which complies with the calculations of the venous capacity and ejection potential derived from the anatomical findings, as mentioned in the previous paragraph. In addition, the results of these measurements showed that the weight- bearing maneuver that was primarily intended to activate the foot pump achieved only 20% of the ejection volume registered in the popliteal vein, whereas 80% of the ejection volume in the popliteal vein accrued from the upper two parts of the calf pump (proximal to the probes measuring the flow in deep lower leg veins). Considering that the foot ejected just 3 ml of venous blood into deep lower leg veins, but the weight-bearing maneuver induced ejection of 6.08 ml registered at the point of measurement in deep lower leg veins, it follows that the lower part of the calf muscles (below the probes) contributed approximately 3 ml to the ejection volume. Since the ejection volume produced by the CMP activity was not less than 60 ml, it is clear that 3 ml ejected by the foot pump is negligible and could be ignored. In the interplay of the calf and foot pump activity, the calf and foot pumps work probably in sequence during normal walking: calf pump (extensor muscles inducing dorsiflexion in the ankle joint) → foot pump (foot in contact with the ground) → calf pump (flexor muscles inducing plantar flexion in the ankle joint). When performing tip-toe maneuvers, the toe stand represents the systole of the CMP; the return to the stand on the heel represents the diastole. However, treading fully on the heel activates the foot pump simultaneously. Thus, in contrast to the gait cycle, during the tip-toe maneuvers, the foot pump’s systolic phase occurs during the calf pump’s diastolic phase, i.e., it works de facto as an interference factor. The effect can be demonstrated on the pressure curves. On the descending arms of the pressure curves, i.e., during the diastole of the calf pump, little spikes were registered, apparently evoked by the systolic input of the foot pump. It is evident that the systolic phase of the foot pump does not affect the systolic/diastolic amplitudes produced by the calf pump; the recordings indicate that the hemodynamic effect of the foot pump is negligible compared with the calf pump. This is well comprehensible because the venous blood volume in the foot is irrelevant compared to the blood volume in the calf. In addition, the foot pump cannot produce a significant diastolic decrease in pressure either in the PTV or GSV. The systolic and diastolic pressure amplitudes in the PTV and GSV appear synchronously and are very similar, indicating that they are produced by the calf pump, both in patients with varicose veins and healthy individuals.

Thirty-six patients with varicose veins were investigated by AVP and DUS. Height-matched and 12 years old controls were used for comparison (13). Patients and controls consented to participate in this study. Twenty-one patients with primary varicose veins (group AI) and 15 patients (group AII) with primary deep venous valve incompetence (DVI) underwent biopsies of the gastro-cnemius muscle during the operation. Adductor biopsies obtained from the same limbs served as the control group (group B), and specimens from control participants without venous disease served as the second control group (group C). All the specimens were investigated by superoxide dismutase (SOD), nitric oxide (NO), Na+/K+-ATPase, Ca2+-ATPase, and lactic acid (LD) determinations. Samples were subjected to light and electron microscopy following hematoxylin and eosin (H & E) staining, special ATPase, and cytochrome oxidase/succinate dehydrogenase (COX/SDH) stains.

Normal muscle architecture was seen following H & E, ATPase, and COX/SDH staining, and normal cell meta-bolism was observed in groups B and C specimens. The gastrocnemius muscle in group A showed pathological changes, including disseminated myofibril atrophy, cell denaturation and necrosis, inflammatory cell infiltration, proliferation, and dilation of interfascicular veins. ATPase staining (pH 9.4) demonstrated the grouping of atrophic fibers, especially type I myofibril grouping, accompanied by moderate to severe atrophy of type II muscle fibers. However, no patient had selective type I fiber atrophy. Enhanced enzymatic activity in single or multiple myofibrils was demonstrated by COX/SDH staining in approximately half of the specimens in group AII. In group AII, electron microscopy showed swelling, myelin figure denaturation of mitochondria, disruption of the myofibrils, and increased lipid droplets in the gastrocnemius muscle. Increased concentration of LD was found in most specimens from patients in group A. There were also reductions of SOD, NO, Na+/K+-ATPase biochemical activity, and Ca2+-ATPase with increasing concentration of LD in these patients, most prominently in group AII. We saw a correlation between AVP assessments and the gastrocnemius muscle’s bioche-mical measurements and morphological appearances.

Myosatellite cells, also known as satellite cells or muscle stem cells (MuSCs), are small multipotent cells with very little cytoplasm in mature muscle. Upon activation, satellite cells can re-enter the cell cycle to proliferate and differentiate into myoblasts. Satellite stem cell activation is the process by which quiescent precursor cells resident on muscle fibers are recruited to cycle and move. Two processes are reported to affect satellite cell activation. Skeletal muscle can regenerate remarkably after injury, a property conferred by a resident population of MuSCs. In response to injury, MuSCs must double their cellular content to divide, a process requiring significant new biomass in the form of nucleotides, phospholipids, and amino acids. This new biomass is derived from a series of intracellular metabolic cycles and alternative carbon routing. Muscle regeneration is complex, requiring the coordinated activity of inflammatory cells, fibroblasts, mesenchymal cells, and MuSCs to ensure the complete restoration of vasculature, nerves, and myofibers (14). As mature myofibers are post-mitotic, muscle regeneration depends on an adequate population of viable MuSCs. In the absence of injury, MuSCs typically exist in a quiescent state outside the cell cycle, residing between the plasma membrane of a myofiber and the basement membrane (15). During homeostasis, MuSCs do not actively proliferate and typically account for 2∼10% of myonuclei, depending on age, sex, and muscle type (16). Upon activation, MuSCs produce a progeny of myogenic cells that can differentiate, culmi-nating in the formation of mature muscle fiber. During this process, MuSCs typically become specified to the myogenic lineage after activation and then undergo multiple rounds of proliferation to generate sufficient myonuclei to support protein synthesis and mature muscle formation (17). These proliferating myogenic precursors (myoblasts) then exit the cell cycle and terminally differentiate into myocytes which fuse to form myotubes. Muscle regeneration is completed through further rounds of myoblast fusion and muscle fiber maturation (18). Importantly, a small subpopulation of myoblasts returns to quiescence to restore the MuSC pool.

Mitochondria are highly dynamic organelles that undergo cycles of fusion and fission important for their function, maintenance, and quality control, as well as the direct or indirect role in different types of stem cells’ fate decisions. Stem cells have the potential for numerous biomedical applications; however, the major bottleneck in the stem cell field is the differentiation and maintenance of stemness of stem cells. The regulatory factor involved in stem cell fate decision and development is unclear. A recent report suggests that mitochondria also play a role in maintaining pluripotency and cell fate decision (19). Mitochondria are considered highly plastic organelles. This plasticity enables the mitochondria to undergo morphological and functional changes in response to cellular demands. Stem cells also need to remain functionally plastic, i.e., to have the ability to “decide” whether to remain quiescent or undergo activation upon signaling cues to support tissue function and homeostasis. Mitochondrial plasticity is thought to enable this reshaping of stem cell functions, integrating signaling cues with stem cell outcomes. Indeed, recent evidence highlights the crucial role of maintaining mitochondrial plasticity for stem cell biology. For example, tricarboxylic acid cycle metabolites generated and metabolized in the mitochondria serve as cofactors for epigenetic enzymes, thereby coupling mitochondrial metabolism and transcrip-tional regulation. Another layer of mitochondrial plasticity has emerged, pointing toward mitochondrial dynamics in regulating stem cell fate decisions. Imposing imbalanced mitochondrial dynamics by manipulating the expression levels of the key molecular regulators of this process influences cellular outcomes by changing the nuclear transcriptional program. Moreover, reactive oxygen species (ROS) have also been shown to play an important role in regulating transcriptional profiles in stem cells. So we focused on recent findings demonstrating that mitochondria are essential regulators of stem cell activation and fate decisions and the suggested mechanisms and alternative routes for mitochondria-to-nucleus communications (*cited from) (20).

Most adult stem cells mainly use aerobic glycolysis for ATP production. This phenomenon in stem cells is known as the “Warburg effect,” which was first described in cancer cells (18). Most cancer cells produce energy through a high rate of glycolysis even when there is sufficient oxygen supply, a phenomenon termed the “Warburg effect.” The precise mechanism of the Warburg effect remains unknown. This phenomenon also came into the spotlight in cellular reprogramming, or induced pluripotent stem cell (iPSC) generation (21), and a metabolic switch from oxidative phosphorylation in mouse embryonic fibroblasts (MEFs) to glycolysis in reprogrammed iPSCs. This phenomenon is commonly observed in various cancers, which display a highly proliferative state. It raises the question of why should proliferating cells choose an inefficient pathway to produce energy? Cell division requires energy and various cellular constituents, such as nucleotides, amino acids, and lipids. Glycolysis and the pentose phosphate pathway can account for cellular constituents and ATP (22). Reducing mitochondrial metabolism may also allow a low level of harmful free radicals, such as ROS. Therefore, glycolysis would benefit the actively proliferating stem cells to self-renew and maintain cell states (23).

Interestingly, the proteins related to mitochondrial dyna-mics, such as fusion and fission, are crucial for pluri-potential reprogramming. During reprogramming (iPSC generation) and re-differentiation of iPSCs, mitochondrial morphology changes dynamically; mitochondria become elongated and globular-shaped after re-differentiation into a neural lineage as the mitochondrial morphology changes dynamically during the process of reprogramming (24-26). Several studies have suggested that mitochondrial dynamics and energy metabolism are critical for reprogramming (27).

Current research has shown that autophagy dysfunction may contribute to the pathogenesis of some myopathies through the impairment of myofibers regeneration. Studies of autophagy inhibition also indicate the importance in muscle regeneration, while activation of autophagy can restore muscle function in some myopathies. The roles of autophagy (organelle/protein degradation, energy facilitation, and/or others) vary at different myogenic stages of the repair process. When the muscle is in homeostasis, basal autophagy can maintain the quiescence state and stemness of MuSCs by renewing organelle and protein. After an injury, the increased autophagy flux contributes to meeting the biological energy demand of MuSCs during activation and proliferation. By mitochondrial remodeling, autophagy during differentiation can promote metabolic transformation and balance mitochondrial-mediated apoptosis signals in myoblasts. Autophagy in mature myofibers is also essential for the degradation of necrotic myofibers and may affect the dynamics of MuSCs by affecting the secretion spectrum of myofibers or the recruitment of supporting cells. Except for myogenic cells, autophagy also plays an important role in regulating the function of non-myogenic cells in the muscle microenvironment, which is also essential for successful muscle recovery. Autophagy can regulate the immune microenvironment during muscle regeneration through the recruitment and polarization of macrophages, while autophagy in endothelial cells can regulate muscle regeneration in an angiogenic or angiogenesis-independent manner. Drug or nutrition-targeted autophagy has been preliminarily shown to restore muscle function in myopathies by promoting muscle regeneration, and further understanding the role and mechanism of autophagy in various cell types during muscle regeneration will enable more effective combinatorial therapeutic strategies. Auto-phagy is an intracellular degradation system that delivers cytoplasmic constituents to the lysosome. Autophagy consists of several sequential steps: sequestration, transport to lysosomes, degradation, and utilization of degradation products. Each step may exert different functions. Although it can also induce cell death, autophagy can be described as a degradation mechanism rather than a form of cell death. Of the cell death types, autophagy has the highest survival superiority, followed by apoptosis, with necrosis having the lowest survival superiority. Autophagy is instinctively induced or inhibited (28-30).

Thus, two or three types of cell death may be induced simultaneously or successively when cells are exposed to certain stimuli. If the three types of cell death are placed on an axis according to their survival superiority, autophagy and necrosis would be placed at opposing ends, whereas apoptosis would be placed in the middle; furthermore, programmed necrosis would be placed between necrosis and apoptosis.

Treatment of varicose veins through muscle regeneration (*A-pretreatment, B-posttreatment) (Fig. 1-8).

Fig. 1. Reticular varicose veins (case 1).

Fig. 2. Spider, reticular varicose veins (case 2).

Fig. 3. Varicose veins (case 3).

Fig. 4. Varicose veins and leg edema (case 4).

Fig. 5. Varicose veins, venous nodes, and leg edema (case 5).

Fig. 6. Varicose veins (stage 4), varicose eczema, and deep trophic ulcer (case 6).

Fig. 7. Foot varicose veins (case 7).

Fig. 8. Bulging hand veins (case 8).

The efficiency of the CMP relies on normally functioning venous valves, powerful contraction of the calf muscle, full ankle joint movement, and normal muscular fasciae. Any malfunction in this system may contribute to calf pump dysfunction, influencing venous hemodynamics and resulting in venous hypertension. Patients with chronic venous disease have venous reflux, weakness of calf muscle strength, and calf pump dysfunction. In normal human skeletal muscle, types I and II muscle fibers are approximately equal in size and are polygonal rather than angular in configuration. The distribution of fiber types resembles that of a checkerboard, with light and dark fibers arranged in an evenly mixed mosaic pattern. Gastrocnemius muscle from patients with venous disease showed changes characterized by denervation and reinnervation, but biopsy specimens from healthy individuals did not show these changes. Electron microscopy showed denaturation of mitochondria and disruption of the myofibrils in the AII group. In muscle cells, mitochondria are scattered among and around the myofibrils, providing the ATP needed to power muscular contractions. Na+/K+-ATPase, found in plasma membranes of most animal cells, catalyzes ATP-dependent transport of Na+ out of a cell in exchange for K+ entering the muscle cell. Ca2+-ATPases, in the endoplasmic reticulum (ER) and plasma membranes of muscle cells, catalyzes ATP-dependent transport of Ca2+ away from the cytosol into the ER lumen or out of the cell. We found reduced ATPase activity in group AII specimens, which might lead to an impaired ability to maintain or restore Na+ and K+ balance across the sarcolemma during repeated muscle contractions. An abnormal intracellular Ca2+ concentration would result in defective muscle func-tion, impacting the muscle excitation-contraction coupling, causing prolonged physiological Ca2+-elevation, slowing of relaxation, and leading to mitochondrial damage and disorganization of myofibrils and muscle weakness (30). This might cause calf muscle fatigue in chronic venous insufficiency patients. It has been shown that failure of Ca2+ release is strongly implicated in muscle fatigue; depressed Na+/K+-ATPase enzyme activity of up to 17% has been reported during fatigue, while training studies have shown a 13∼16% increase in Na+/K+-ATPase activity. SOD is one of the most important and powerful antioxidants in tissue. Contractions of skeletal muscles produce an increased concentration of superoxide anions and activity of hydroxyl radicals in the extracellular space, which alters the skeletal muscle oxidative capacity. Some studies indicated that superoxide radicals play an important role in the pathogenesis of varicose veins, and SOD can inhibit the expression of free radicals and adhesion molecules, protecting skeletal muscle from impairment. We found that biopsies from the AII group had low levels of muscle SOD compared to the other groups. It has been demonstrated that interval and continuous exercise training increases SOD activity, and recovery time after exercise was shortened when Vitamins C or E or exogenous SOD was supplied. It may be possible to use this type of treatment in patients with venous diseases to improve CMP function. The concentration of lactic acid in the blood (LAB) is one of the main measures used to evaluate skeletal muscle fatigue in sports medicine, and it is mainly influenced by the balance between the production and elimination of lactic acid in skeletal muscle (LAM). The concentration of LAM directly evaluates LD metabolism in muscle. We measured the concentration of LAM to estimate the metabolism of gastrocnemius cells and found that the concentration of LAM in group A including popliteal vein valves and deep venous incompetence patients, was increased compared to control specimens. This would lower the intracellular pH and alter the functional characteristics of key enzymes in muscle cells. LD diffuses out of the muscle fibers and enters the Cori cycle, where LD is metabolized in the liver returning glucose to the muscle cells during recovery. Until LAM levels fall, premature muscle fatigue may be the result. A review of the literature suggests that the effects of NO on skeletal muscle fibers can be classified into direct and cGMP-mediated effects. These include NO-stimulated glucose uptake, glycolysis, and mitochondrial respiration, increasing the shortening velocity of loaded or unloaded contractions. NO has a clear role in regulating basal vascular tone at rest and contributing partly to the blood flow in recovery after exhaustive exercise. Impaired biochemical function and morphology have also been reported in samples of skeletal muscle from patients with the arterial occlusive disease or disuse atrophy. Ischemia affects each muscle fiber type differently according to its particular metabolic and functional properties. In some studies, effects on type I fibers predominated, and others indicated a selective vulnerability of the fast glycolytic fibers. Studies by Clyne et al. showed decreased levels of aerobic enzymes paralleling decreased Doppler ankle pressure, while claudicants demonstrated increased levels of anaerobic enzymes. We consider that anoxia due to arterial ischemia or venous congestion would lead to skeletal muscle impairment characterized by denervation and reinnervation.

We found structural and metabolic abnormalities in the calf skeletal muscle cells of patients with venous insufficiency, which were most prominent in the most severely affected patients. Some of the symptoms reported by patients in their lower limbs may be attributable to these changes. It has been suggested that muscle regeneration may be very useful in addressing similar muscle changes reported in patients with peripheral arterial diseases. Additional therapies, including physical therapy or thera-peutic exercise, muscle nutrition, and antioxidant therapy, might improve muscle regeneration of venous disease.

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Review Article

Ann Phlebology 2022; 20(2): 68-77

Published online December 31, 2022 https://doi.org/10.37923/phle.2022.20.2.68

Copyright © Annals of phlebology.

New Treatment of Varicose Veins through Muscle Regeneration of Lower Leg Muscles (Especially Calf Muscle) Without Removal of Varicose Veins

Ki Ji Lee, M.D., Ph.D.

Department of Pediatrics, Woorisoa Hospital, Seoul, Korea

Correspondence to:Ki Ji Lee, 15 Saemal-ro, Guro-gu, Seoul 47848, Korea, Deparment of Pediatrics, Woorisoa Seoul Hospital
Tel: 02-858-0100, Fax: 02-856-0032
E-mail: leekiji@naver.com

Abstract

The calf muscle pump is the motive force enhancing venous blood return from the lower extremity to the heart. It causes displacement of venous blood in both vertical and horizontal directions, generates ambulatory pressure gradient between the thigh and lower leg veins, and bidirectional streaming within calf perforators. Ambulatory pressure gradient triggers venous reflux in incompetent veins, inducing ambulatory venous hypertension in the lower leg and foot. Bidirectional flow in calf perforators enables quick pressure equalization between deep and superficial veins of the lower leg; the outward (into the superficial veins) oriented component of the bidirectional flow taking place during calf muscle contraction is not a pathological reflux but a physiological centripetal flow streaming via the great saphenous vein into the femoral vein. Calf perforators are communicating channels between both systems, making them conjoined vessels; they are not involved in generating pathological hemodynamic situations and do not cause ambulatory venous hypertension. Pressure gradient arising during calf pump activity between the femoral vein and the saphenous remnant after abolishing saphenous reflux triggers biophysical and biochemical events, which might induce recurrence. Thus, abolishing saphenous reflux removes the hemodynamic disturbance but simultaneously generates a precondition for reflux recurrence and the return of the previous pathological situation; this chain of events has been termed the hemodynamic paradox. But this review showed that varicose veins could be improved quickly through lower leg muscles (especially calf muscle) regeneration by increasing mitochondrial cellular energy (adenosine triphosphate) of leg muscles without removing varicose veins.

Keywords: Calf muscle pump, Venous hemodynamics, Muscle regeneration, Muscle stem cells (satellite cells), Mitochondrial energy (adenosine triphosphate)

INTRODUCTION

Vis a tergo, the remainder of the cardiac energy propelling venous blood from the lower extremities against the gravitation toward the heart is not powerful enough to move the venous blood toward the heart effectively; the pressure difference between the venules and the right atrium is approximately 20 mmHg. In addition to this motive force, the calf muscle pump (CMP) constitutes an effective mechanism that promotes the drainage of venous blood from the lower extremity, produces a marked increase of systolic pressure, decreases diastolic pressure in deep and superficial veins of the lower leg, and entails a distinct physiological decrease of ambulatory venous pressure (AVP) in the lower leg veins. The requirements for efficiency and effectiveness of the CMP are patency and perfect competence of deep lower leg veins, no impedi-ments in the venous outflow tract, and undisturbed contractility of the calf musculature. The venous reflux compromises the physiological effectivity of the CMP since the venous blood volume expelled by the CMP is more or less replaced by the refluxing volume. The contribution of the foot pump to the venous return from the lower extremity remains clear. This review article intends to evaluate the physiological significance of the CMP and the foot pump, assess the systolic and diastolic pressure behavior in deep and superficial veins of the lower leg during calf pump activity, and evaluate the flow behavior in calf perforators resulting from systolic and diastolic pressure differences.

ANATOMY AND FUNCTION OF THE CMP

The calf musculature and the deep lower leg veins form the CMP. The calf musculature consists of

- 1. The posterior compartment inducing plantar flexion of the ankle joint and the toes, as well as supination of the foot.

- 2. The anterior compartment inducing dorsal flexion of the ankle joint and the toes.

- 3. The lateral compartment inducing plantar flexion of the ankle joint and pronation of the foot.

Deep lower leg veins consist of three paired conductive veins: the posterior tibial vein (PTV), anterior tibial vein, peroneal vein, and the muscular veins contained in the sural musculature (most important: gastrocnemius and soleus veins). The CMP is called “peripheral heart”: contractions of the sural muscles eject the venous blood centripetally toward the heart and promote venous blood return from the lower extremity against the gravitation force substantially. During calf muscle contraction, the pressure rises conside-rably in deep lower leg veins (on average of 75 mmHg) and a little less in the great saphenous vein (GSV). The systolic pressure difference between the PTV and GSV amounts to approximately 40 mmHg in healthy individuals and merely 14 mmHg in patients with varicose veins (1-3); the unequal flow rate capacity of the calf perforators in healthy individuals and patients with varicose vein causes this pressure gradient difference. Thus, during calf muscle contractions, the venous blood is ejected mainly into the popliteal vein, but a smaller part escapes through calf perforators into the GSV and streams further in the centripetal direction toward the heart. During calf muscle relaxation, the diastolic pressure decreases in deep lower leg veins and GSV; the pressure gradient reverses, and the diastolic pressure in the GSV is approximately 13 mmHg higher than in the PTV (2,3). The systolic and diastolic pressure changes are produced in deep veins. Pressure transmissions between the PTV and GSV occur through the connecting channels, through calf perforators, documented by simultaneous pressure recordings. The pressure trans-missions could not occur without bidirectional streaming through calf perforators, i.e., without systolic outward and diastolic inward flows. The outward flow produces a steep systolic increase in pressure in the GSV through calf perforators. This flow is centripetal, i.e., physiological; it continues via GSV toward to heart and can be assessed in the GSV by duplex ultrasonography (DUS). In healthy individuals with competent calf perforators, there was a steep increase in systolic pressure in the GSV; however, a similar steep increase in the PTV could not occur. Similar conditions exist in the heart across the competent and incompetent mitral valve. A competent mitral valve precludes a systolic increase in pressure in the left atrium. Regurgitation through an incompetent mitral valve induces increased pressure in the left atrium. Thus, with some benevolence, CMP can be compared with the left ventricle and the saphenous system with the left atrium. The pressure in the popliteal and femoral vein oscillates during the CMP activity but does not decrease below the hydrostatic pressure value during the diastolic phase (1,2,4). The pressure difference arising in this way between the popliteal/femoral veins and deep lower leg veins has been called ambulatory pressure gradient; its value is 37.4±6.4 mmHg (5). Ambulatory pressure gradient triggers reflux in incompetent veins connecting both poles of the pressure gradient. The performance of the CMP depends primarily on the patency and impeccable competence of the deep conductive veins of the lower leg (posterior and anterior tibial veins and peroneal veins). The incompetence of these veins disturbs the performance of the CMP. Further factors impairing CMP performance are obstacles in the outflow venous tracts and reduced calf muscle strength due to muscle weakness, atrophy, or palsy. Plethysmographic findings enable quantification of the CMP performance. Ejection volume produced by the CMP is an important parameter indicating its effectivity; it is determined by air-plethysmography; the normal values range from 60 to 90 ml (5). Ejection fraction is another parameter used for evaluating CMP performance; it is expressed in percentage and is calculated as the volume of blood ejected with one calf muscle contraction divided by the functional venous volume. Since this parameter depends on the value of the functional venous volume, it is not suitable for evaluating results after therapeutic procedures that reduce the venous volume contained in varicose veins. The ejection fraction, i.e., the quotient EV/VV, improves and simulates improved calf muscle performance, but the pumping effectivity remains unaffected. Other valuable plethysmographic para-meters registered after a series of calf muscle contractions and relaxations, such as refill time t-50 and t-90, refill volume gained by strain gauge plethysmography, and residual volume gained by air plethysmography, are influenced by the CMP performance and venous reflux. Saphenous reflux simulates CMP malfunction because the refluxing volume devaluates the ejection volume. Therefore, the last mentioned parameters can be considered for evaluating the CMP performance only after eliminating saphenous reflux. As a rule, in patients with primary varicose veins, even the severest hemodynamic disorders in clinical, etiological, anatomical, and pathological classes C4-C6 are caused by saphenous reflux, whereas the performance of the CMP remains unaffected. The elimina-tion of saphenous reflux restores physiological AVP conditions and normal plethysmographic values, which would be impossible if CMP performance were impaired (6-9).

ANATOMY AND FUNCTION OF THE FOOT PUMP

The plantar venous plexus forms a substantial part of the venous reservoir of the foot pump. According to Binns and Pho (10), the plantar venous plexus consists, in addition to subtle veins, mainly of the large lateral plantar vein that is partly double-barreled, of a little smaller medial plantar vein and the deep plantar venous arch. The plantar veins are convoluted and fitted with valves directing the flow into the PTV. From the diameter and length of these veins, it can be computed that the plantar venous plexus contains approximately 6 ml of blood. In another study, White et al. (11) reported an average of 2.7 large veins in the plantar venous plexus with a diameter of 4.0±1.0 mm. The entire length of these veins along the forefoot was approximately 30 cm, meaning they contained approximately 5 ml of blood. So, the total venous volume of the sole, including the blood volume in the foot muscles, might hardly reach 10 ml. The contribution of the foot pump to the venous return is still disputable. Plantar veins empty due to compression exerted by weight bearing. Due to the foot’s anatomical configuration, only the lateral part is involved in the pumping process. When the normal ejection fraction of 60% is considered, the estimated ejection volume of the foot pump would be approximately 3∼4 ml; the large part of this very small volume is ejected into deep lower leg veins, and a little part might be ejected into the GSV. Broderick et al. (12) researched venous emptying from the foot using several provoking maneuvers: weight bearing, toe curl, electrical stimulation of the tibial nerve at the medial malleolus, and passive compression. Flow measurements in the particular lower leg veins (posterior tibial, anterior tibial, and peroneal veins) and popliteal veins were performed using DUS; the flow in the GSV was not measured. Flow measurements in the deep lower leg veins were performed at the level between the lower and middle part of the lower leg. Values of the ejection volume were obtained by multiplying the cross-sectional area by the mean velocity and total flow duration. Electrical stimulation of the foot muscles ejected just 1 ml of venous blood into the lower leg veins altogether, indicating that the contraction of the metatarsal muscles exerts just a little effect. Intermittent pneumatic foot compression produced a venous outflow of 2.18 ml, which is likewise negligible. Weight-bearing maneuver expelled 6.08 ml and toe curls 5.03 ml of venous blood into deep lower leg veins. Oddly enough, weight-bearing maneuvers and toe curls induced in the popliteal vein a remarkable centripetal flow of 33.34 ml and 31.04 ml, respectively, showing that weight-bearing maneuvers and toe curls activated the foot pump and, more effectively also, the CMP. It is quite difficult to selectively provoke the foot pump activity without evoking in the weigh-bearing limb involuntary contractions of the calf musculature that occur to ensure the balance of the body. In addition, toe curls activate the CMP (musculus flexor hallucis longus and musculus flexor digitorum longus). Since the flow measurements in the lower leg veins were performed at the level between the lower and middle third of the lower leg, it can be deduced that the upper section of the calf musculature where the substantial mass of calf musculature is concentrated might be the most important part of the CMP. The findings documented that intermittent pneumatic compression of the sole and active contractions of the foot muscles could not expel a significant volume of venous blood from the foot. The total volume ejected by these two maneuvers into deep lower leg veins amounted to 3.18 ml (1 ml due to electrical stimulation of the tibial nerve, 2.18 ml due to intermittent pneumatic compression), which complies with the calculations of the venous capacity and ejection potential derived from the anatomical findings, as mentioned in the previous paragraph. In addition, the results of these measurements showed that the weight- bearing maneuver that was primarily intended to activate the foot pump achieved only 20% of the ejection volume registered in the popliteal vein, whereas 80% of the ejection volume in the popliteal vein accrued from the upper two parts of the calf pump (proximal to the probes measuring the flow in deep lower leg veins). Considering that the foot ejected just 3 ml of venous blood into deep lower leg veins, but the weight-bearing maneuver induced ejection of 6.08 ml registered at the point of measurement in deep lower leg veins, it follows that the lower part of the calf muscles (below the probes) contributed approximately 3 ml to the ejection volume. Since the ejection volume produced by the CMP activity was not less than 60 ml, it is clear that 3 ml ejected by the foot pump is negligible and could be ignored. In the interplay of the calf and foot pump activity, the calf and foot pumps work probably in sequence during normal walking: calf pump (extensor muscles inducing dorsiflexion in the ankle joint) → foot pump (foot in contact with the ground) → calf pump (flexor muscles inducing plantar flexion in the ankle joint). When performing tip-toe maneuvers, the toe stand represents the systole of the CMP; the return to the stand on the heel represents the diastole. However, treading fully on the heel activates the foot pump simultaneously. Thus, in contrast to the gait cycle, during the tip-toe maneuvers, the foot pump’s systolic phase occurs during the calf pump’s diastolic phase, i.e., it works de facto as an interference factor. The effect can be demonstrated on the pressure curves. On the descending arms of the pressure curves, i.e., during the diastole of the calf pump, little spikes were registered, apparently evoked by the systolic input of the foot pump. It is evident that the systolic phase of the foot pump does not affect the systolic/diastolic amplitudes produced by the calf pump; the recordings indicate that the hemodynamic effect of the foot pump is negligible compared with the calf pump. This is well comprehensible because the venous blood volume in the foot is irrelevant compared to the blood volume in the calf. In addition, the foot pump cannot produce a significant diastolic decrease in pressure either in the PTV or GSV. The systolic and diastolic pressure amplitudes in the PTV and GSV appear synchronously and are very similar, indicating that they are produced by the calf pump, both in patients with varicose veins and healthy individuals.

Thirty-six patients with varicose veins were investigated by AVP and DUS. Height-matched and 12 years old controls were used for comparison (13). Patients and controls consented to participate in this study. Twenty-one patients with primary varicose veins (group AI) and 15 patients (group AII) with primary deep venous valve incompetence (DVI) underwent biopsies of the gastro-cnemius muscle during the operation. Adductor biopsies obtained from the same limbs served as the control group (group B), and specimens from control participants without venous disease served as the second control group (group C). All the specimens were investigated by superoxide dismutase (SOD), nitric oxide (NO), Na+/K+-ATPase, Ca2+-ATPase, and lactic acid (LD) determinations. Samples were subjected to light and electron microscopy following hematoxylin and eosin (H & E) staining, special ATPase, and cytochrome oxidase/succinate dehydrogenase (COX/SDH) stains.

Normal muscle architecture was seen following H & E, ATPase, and COX/SDH staining, and normal cell meta-bolism was observed in groups B and C specimens. The gastrocnemius muscle in group A showed pathological changes, including disseminated myofibril atrophy, cell denaturation and necrosis, inflammatory cell infiltration, proliferation, and dilation of interfascicular veins. ATPase staining (pH 9.4) demonstrated the grouping of atrophic fibers, especially type I myofibril grouping, accompanied by moderate to severe atrophy of type II muscle fibers. However, no patient had selective type I fiber atrophy. Enhanced enzymatic activity in single or multiple myofibrils was demonstrated by COX/SDH staining in approximately half of the specimens in group AII. In group AII, electron microscopy showed swelling, myelin figure denaturation of mitochondria, disruption of the myofibrils, and increased lipid droplets in the gastrocnemius muscle. Increased concentration of LD was found in most specimens from patients in group A. There were also reductions of SOD, NO, Na+/K+-ATPase biochemical activity, and Ca2+-ATPase with increasing concentration of LD in these patients, most prominently in group AII. We saw a correlation between AVP assessments and the gastrocnemius muscle’s bioche-mical measurements and morphological appearances.

MUSCLE STEM CELL AND MUSCLE REGENERATION

Myosatellite cells, also known as satellite cells or muscle stem cells (MuSCs), are small multipotent cells with very little cytoplasm in mature muscle. Upon activation, satellite cells can re-enter the cell cycle to proliferate and differentiate into myoblasts. Satellite stem cell activation is the process by which quiescent precursor cells resident on muscle fibers are recruited to cycle and move. Two processes are reported to affect satellite cell activation. Skeletal muscle can regenerate remarkably after injury, a property conferred by a resident population of MuSCs. In response to injury, MuSCs must double their cellular content to divide, a process requiring significant new biomass in the form of nucleotides, phospholipids, and amino acids. This new biomass is derived from a series of intracellular metabolic cycles and alternative carbon routing. Muscle regeneration is complex, requiring the coordinated activity of inflammatory cells, fibroblasts, mesenchymal cells, and MuSCs to ensure the complete restoration of vasculature, nerves, and myofibers (14). As mature myofibers are post-mitotic, muscle regeneration depends on an adequate population of viable MuSCs. In the absence of injury, MuSCs typically exist in a quiescent state outside the cell cycle, residing between the plasma membrane of a myofiber and the basement membrane (15). During homeostasis, MuSCs do not actively proliferate and typically account for 2∼10% of myonuclei, depending on age, sex, and muscle type (16). Upon activation, MuSCs produce a progeny of myogenic cells that can differentiate, culmi-nating in the formation of mature muscle fiber. During this process, MuSCs typically become specified to the myogenic lineage after activation and then undergo multiple rounds of proliferation to generate sufficient myonuclei to support protein synthesis and mature muscle formation (17). These proliferating myogenic precursors (myoblasts) then exit the cell cycle and terminally differentiate into myocytes which fuse to form myotubes. Muscle regeneration is completed through further rounds of myoblast fusion and muscle fiber maturation (18). Importantly, a small subpopulation of myoblasts returns to quiescence to restore the MuSC pool.

MITOCHONDRIAL PLASTICITY IN CELL FATE REGULATION

Mitochondria are highly dynamic organelles that undergo cycles of fusion and fission important for their function, maintenance, and quality control, as well as the direct or indirect role in different types of stem cells’ fate decisions. Stem cells have the potential for numerous biomedical applications; however, the major bottleneck in the stem cell field is the differentiation and maintenance of stemness of stem cells. The regulatory factor involved in stem cell fate decision and development is unclear. A recent report suggests that mitochondria also play a role in maintaining pluripotency and cell fate decision (19). Mitochondria are considered highly plastic organelles. This plasticity enables the mitochondria to undergo morphological and functional changes in response to cellular demands. Stem cells also need to remain functionally plastic, i.e., to have the ability to “decide” whether to remain quiescent or undergo activation upon signaling cues to support tissue function and homeostasis. Mitochondrial plasticity is thought to enable this reshaping of stem cell functions, integrating signaling cues with stem cell outcomes. Indeed, recent evidence highlights the crucial role of maintaining mitochondrial plasticity for stem cell biology. For example, tricarboxylic acid cycle metabolites generated and metabolized in the mitochondria serve as cofactors for epigenetic enzymes, thereby coupling mitochondrial metabolism and transcrip-tional regulation. Another layer of mitochondrial plasticity has emerged, pointing toward mitochondrial dynamics in regulating stem cell fate decisions. Imposing imbalanced mitochondrial dynamics by manipulating the expression levels of the key molecular regulators of this process influences cellular outcomes by changing the nuclear transcriptional program. Moreover, reactive oxygen species (ROS) have also been shown to play an important role in regulating transcriptional profiles in stem cells. So we focused on recent findings demonstrating that mitochondria are essential regulators of stem cell activation and fate decisions and the suggested mechanisms and alternative routes for mitochondria-to-nucleus communications (*cited from) (20).

WARBURG EFFECT: SURVIVAL STRATEGY OF PROLIFERATIVE GLYCOLYTIC CELLS

Most adult stem cells mainly use aerobic glycolysis for ATP production. This phenomenon in stem cells is known as the “Warburg effect,” which was first described in cancer cells (18). Most cancer cells produce energy through a high rate of glycolysis even when there is sufficient oxygen supply, a phenomenon termed the “Warburg effect.” The precise mechanism of the Warburg effect remains unknown. This phenomenon also came into the spotlight in cellular reprogramming, or induced pluripotent stem cell (iPSC) generation (21), and a metabolic switch from oxidative phosphorylation in mouse embryonic fibroblasts (MEFs) to glycolysis in reprogrammed iPSCs. This phenomenon is commonly observed in various cancers, which display a highly proliferative state. It raises the question of why should proliferating cells choose an inefficient pathway to produce energy? Cell division requires energy and various cellular constituents, such as nucleotides, amino acids, and lipids. Glycolysis and the pentose phosphate pathway can account for cellular constituents and ATP (22). Reducing mitochondrial metabolism may also allow a low level of harmful free radicals, such as ROS. Therefore, glycolysis would benefit the actively proliferating stem cells to self-renew and maintain cell states (23).

MITOCHONDRIAL DYNAMICS IN THE REPROGRAMMING PROCESS

Interestingly, the proteins related to mitochondrial dyna-mics, such as fusion and fission, are crucial for pluri-potential reprogramming. During reprogramming (iPSC generation) and re-differentiation of iPSCs, mitochondrial morphology changes dynamically; mitochondria become elongated and globular-shaped after re-differentiation into a neural lineage as the mitochondrial morphology changes dynamically during the process of reprogramming (24-26). Several studies have suggested that mitochondrial dynamics and energy metabolism are critical for reprogramming (27).

AUTOPHAGY, MITOCHONDRIA, AND TISSUE REGENERATION

Current research has shown that autophagy dysfunction may contribute to the pathogenesis of some myopathies through the impairment of myofibers regeneration. Studies of autophagy inhibition also indicate the importance in muscle regeneration, while activation of autophagy can restore muscle function in some myopathies. The roles of autophagy (organelle/protein degradation, energy facilitation, and/or others) vary at different myogenic stages of the repair process. When the muscle is in homeostasis, basal autophagy can maintain the quiescence state and stemness of MuSCs by renewing organelle and protein. After an injury, the increased autophagy flux contributes to meeting the biological energy demand of MuSCs during activation and proliferation. By mitochondrial remodeling, autophagy during differentiation can promote metabolic transformation and balance mitochondrial-mediated apoptosis signals in myoblasts. Autophagy in mature myofibers is also essential for the degradation of necrotic myofibers and may affect the dynamics of MuSCs by affecting the secretion spectrum of myofibers or the recruitment of supporting cells. Except for myogenic cells, autophagy also plays an important role in regulating the function of non-myogenic cells in the muscle microenvironment, which is also essential for successful muscle recovery. Autophagy can regulate the immune microenvironment during muscle regeneration through the recruitment and polarization of macrophages, while autophagy in endothelial cells can regulate muscle regeneration in an angiogenic or angiogenesis-independent manner. Drug or nutrition-targeted autophagy has been preliminarily shown to restore muscle function in myopathies by promoting muscle regeneration, and further understanding the role and mechanism of autophagy in various cell types during muscle regeneration will enable more effective combinatorial therapeutic strategies. Auto-phagy is an intracellular degradation system that delivers cytoplasmic constituents to the lysosome. Autophagy consists of several sequential steps: sequestration, transport to lysosomes, degradation, and utilization of degradation products. Each step may exert different functions. Although it can also induce cell death, autophagy can be described as a degradation mechanism rather than a form of cell death. Of the cell death types, autophagy has the highest survival superiority, followed by apoptosis, with necrosis having the lowest survival superiority. Autophagy is instinctively induced or inhibited (28-30).

Thus, two or three types of cell death may be induced simultaneously or successively when cells are exposed to certain stimuli. If the three types of cell death are placed on an axis according to their survival superiority, autophagy and necrosis would be placed at opposing ends, whereas apoptosis would be placed in the middle; furthermore, programmed necrosis would be placed between necrosis and apoptosis.

Treatment of varicose veins through muscle regeneration (*A-pretreatment, B-posttreatment) (Fig. 1-8).

Figure 1. Reticular varicose veins (case 1).

Figure 2. Spider, reticular varicose veins (case 2).

Figure 3. Varicose veins (case 3).

Figure 4. Varicose veins and leg edema (case 4).

Figure 5. Varicose veins, venous nodes, and leg edema (case 5).

Figure 6. Varicose veins (stage 4), varicose eczema, and deep trophic ulcer (case 6).

Figure 7. Foot varicose veins (case 7).

Figure 8. Bulging hand veins (case 8).

DISCUSSION

The efficiency of the CMP relies on normally functioning venous valves, powerful contraction of the calf muscle, full ankle joint movement, and normal muscular fasciae. Any malfunction in this system may contribute to calf pump dysfunction, influencing venous hemodynamics and resulting in venous hypertension. Patients with chronic venous disease have venous reflux, weakness of calf muscle strength, and calf pump dysfunction. In normal human skeletal muscle, types I and II muscle fibers are approximately equal in size and are polygonal rather than angular in configuration. The distribution of fiber types resembles that of a checkerboard, with light and dark fibers arranged in an evenly mixed mosaic pattern. Gastrocnemius muscle from patients with venous disease showed changes characterized by denervation and reinnervation, but biopsy specimens from healthy individuals did not show these changes. Electron microscopy showed denaturation of mitochondria and disruption of the myofibrils in the AII group. In muscle cells, mitochondria are scattered among and around the myofibrils, providing the ATP needed to power muscular contractions. Na+/K+-ATPase, found in plasma membranes of most animal cells, catalyzes ATP-dependent transport of Na+ out of a cell in exchange for K+ entering the muscle cell. Ca2+-ATPases, in the endoplasmic reticulum (ER) and plasma membranes of muscle cells, catalyzes ATP-dependent transport of Ca2+ away from the cytosol into the ER lumen or out of the cell. We found reduced ATPase activity in group AII specimens, which might lead to an impaired ability to maintain or restore Na+ and K+ balance across the sarcolemma during repeated muscle contractions. An abnormal intracellular Ca2+ concentration would result in defective muscle func-tion, impacting the muscle excitation-contraction coupling, causing prolonged physiological Ca2+-elevation, slowing of relaxation, and leading to mitochondrial damage and disorganization of myofibrils and muscle weakness (30). This might cause calf muscle fatigue in chronic venous insufficiency patients. It has been shown that failure of Ca2+ release is strongly implicated in muscle fatigue; depressed Na+/K+-ATPase enzyme activity of up to 17% has been reported during fatigue, while training studies have shown a 13∼16% increase in Na+/K+-ATPase activity. SOD is one of the most important and powerful antioxidants in tissue. Contractions of skeletal muscles produce an increased concentration of superoxide anions and activity of hydroxyl radicals in the extracellular space, which alters the skeletal muscle oxidative capacity. Some studies indicated that superoxide radicals play an important role in the pathogenesis of varicose veins, and SOD can inhibit the expression of free radicals and adhesion molecules, protecting skeletal muscle from impairment. We found that biopsies from the AII group had low levels of muscle SOD compared to the other groups. It has been demonstrated that interval and continuous exercise training increases SOD activity, and recovery time after exercise was shortened when Vitamins C or E or exogenous SOD was supplied. It may be possible to use this type of treatment in patients with venous diseases to improve CMP function. The concentration of lactic acid in the blood (LAB) is one of the main measures used to evaluate skeletal muscle fatigue in sports medicine, and it is mainly influenced by the balance between the production and elimination of lactic acid in skeletal muscle (LAM). The concentration of LAM directly evaluates LD metabolism in muscle. We measured the concentration of LAM to estimate the metabolism of gastrocnemius cells and found that the concentration of LAM in group A including popliteal vein valves and deep venous incompetence patients, was increased compared to control specimens. This would lower the intracellular pH and alter the functional characteristics of key enzymes in muscle cells. LD diffuses out of the muscle fibers and enters the Cori cycle, where LD is metabolized in the liver returning glucose to the muscle cells during recovery. Until LAM levels fall, premature muscle fatigue may be the result. A review of the literature suggests that the effects of NO on skeletal muscle fibers can be classified into direct and cGMP-mediated effects. These include NO-stimulated glucose uptake, glycolysis, and mitochondrial respiration, increasing the shortening velocity of loaded or unloaded contractions. NO has a clear role in regulating basal vascular tone at rest and contributing partly to the blood flow in recovery after exhaustive exercise. Impaired biochemical function and morphology have also been reported in samples of skeletal muscle from patients with the arterial occlusive disease or disuse atrophy. Ischemia affects each muscle fiber type differently according to its particular metabolic and functional properties. In some studies, effects on type I fibers predominated, and others indicated a selective vulnerability of the fast glycolytic fibers. Studies by Clyne et al. showed decreased levels of aerobic enzymes paralleling decreased Doppler ankle pressure, while claudicants demonstrated increased levels of anaerobic enzymes. We consider that anoxia due to arterial ischemia or venous congestion would lead to skeletal muscle impairment characterized by denervation and reinnervation.

CONCLUSION

We found structural and metabolic abnormalities in the calf skeletal muscle cells of patients with venous insufficiency, which were most prominent in the most severely affected patients. Some of the symptoms reported by patients in their lower limbs may be attributable to these changes. It has been suggested that muscle regeneration may be very useful in addressing similar muscle changes reported in patients with peripheral arterial diseases. Additional therapies, including physical therapy or thera-peutic exercise, muscle nutrition, and antioxidant therapy, might improve muscle regeneration of venous disease.

Fig 1.

Figure 1.Reticular varicose veins (case 1).
Annals of Phlebology 2022; 20: 68-77https://doi.org/10.37923/phle.2022.20.2.68

Fig 2.

Figure 2.Spider, reticular varicose veins (case 2).
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Fig 3.

Figure 3.Varicose veins (case 3).
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Fig 4.

Figure 4.Varicose veins and leg edema (case 4).
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Fig 5.

Figure 5.Varicose veins, venous nodes, and leg edema (case 5).
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Fig 6.

Figure 6.Varicose veins (stage 4), varicose eczema, and deep trophic ulcer (case 6).
Annals of Phlebology 2022; 20: 68-77https://doi.org/10.37923/phle.2022.20.2.68

Fig 7.

Figure 7.Foot varicose veins (case 7).
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Fig 8.

Figure 8.Bulging hand veins (case 8).
Annals of Phlebology 2022; 20: 68-77https://doi.org/10.37923/phle.2022.20.2.68

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Vol.21 No.2 Dec 31, 2023, pp. 53~98

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Annals of Phlebology