Leg immobilization and subsequent recovery resistance training affect skeletal muscle angiogenesis related markers in young healthy adults regardless of prior resistance training experience

Leg immobilization and subsequent recovery resistance training affect skeletal muscle angiogenesis related markers in young healthy adults regardless of prior resistance training experience | bioRxiv /* */ /* */ <!-- <!-- /*! * yepnope1.5.4 * (c) WTFPL, GPLv2 */ (function(a,b,c){function d(a){return"[object Function]"==o.call(a)}function e(a){return"string"==typeof a}function f(){}function g(a){return!a||"loaded"==a||"complete"==a||"uninitialized"==a}function h(){var a=p.shift();q=1,a?a.t?m(function(){("c"==a.t?B.injectCss:B.injectJs)(a.s,0,a.a,a.x,a.e,1)},0):(a(),h()):q=0}function i(a,c,d,e,f,i,j){function k(b){if(!o&&g(l.readyState)&&(u.r=o=1,!q&&h(),l.onload=l.onreadystatechange=null,b)){"img"!=a&&m(function(){t.removeChild(l)},50);for(var d in y[c])y[c].hasOwnProperty(d)&&y[c][d].onload()}}var j=j||B.errorTimeout,l=b.createElement(a),o=0,r=0,u={t:d,s:c,e:f,a:i,x:j};1===y[c]&&(r=1,y[c]=[]),"object"==a?l.data=c:(l.src=c,l.type=a),l.width=l.height="0",l.onerror=l.onload=l.onreadystatechange=function(){k.call(this,r)},p.splice(e,0,u),"img"!=a&&(r||2===y[c]?(t.insertBefore(l,s?null:n),m(k,j)):y[c].push(l))}function j(a,b,c,d,f){return q=0,b=b||"j",e(a)?i("c"==b?v:u,a,b,this.i++,c,d,f):(p.splice(this.i++,0,a),1==p.length&&h()),this}function k(){var a=B;return a.loader={load:j,i:0},a}var l=b.documentElement,m=a.setTimeout,n=b.getElementsByTagName("script")[0],o={}.toString,p=[],q=0,r="MozAppearance"in l.style,s=r&&!!b.createRange().compareNode,t=s?l:n.parentNode,l=a.opera&&"[object Opera]"==o.call(a.opera),l=!!b.attachEvent&&!l,u=r?"object":l?"script":"img",v=l?"script":u,w=Array.isArray||function(a){return"[object Array]"==o.call(a)},x=[],y={},z={timeout:function(a,b){return b.length&&(a.timeout=b[0]),a}},A,B;B=function(a){function b(a){var a=a.split("!"),b=x.length,c=a.pop(),d=a.length,c={url:c,origUrl:c,prefixes:a},e,f,g;for(f=0;f<d;f++)g=a[f].split("="),(e=z[g.shift()])&&(c=e(c,g));for(f=0;f<b;f++)c=x[f](c);return c}function g(a,e,f,g,h){var i=b(a),j=i.autoCallback;i.url.split(".").pop().split("?").shift(),i.bypass||(e&&(e=d(e)?e:e[a]||e[g]||e[a.split("/").pop().split("?")[0]]),i.instead?i.instead(a,e,f,g,h):(y[i.url]?i.noexec=!0:y[i.url]=1,f.load(i.url,i.forceCSS||!i.forceJS&&"css"==i.url.split(".").pop().split("?").shift()?"c":c,i.noexec,i.attrs,i.timeout),(d(e)||d(j))&&f.load(function(){k(),e&&e(i.origUrl,h,g),j&&j(i.origUrl,h,g),y[i.url]=2})))}function h(a,b){function c(a,c){if(a){if(e(a))c||(j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}),g(a,j,b,0,h);else if(Object(a)===a)for(n in m=function(){var b=0,c;for(c in a)a.hasOwnProperty(c)&&b++;return b}(),a)a.hasOwnProperty(n)&&(!c&&!--m&&(d(j)?j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}:j[n]=function(a){return function(){var b=[].slice.call(arguments);a&&a.apply(this,b),l()}}(k[n])),g(a[n],j,b,n,h))}else!c&&l()}var h=!!a.test,i=a.load||a.both,j=a.callback||f,k=j,l=a.complete||f,m,n;c(h?a.yep:a.nope,!!i),i&&c(i)}var i,j,l=this.yepnope.loader;if(e(a))g(a,0,l,0);else if(w(a))for(i=0;i (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0];var j=d.createElement(s);var dl=l!='dataLayer'?'&l='+l:'';j.src='//www.googletagmanager.com/gtm.js?id='+i+dl;j.type='text/javascript';j.async=true;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-M677548'); Skip to main content Home About Submit ALERTS / RSS Search for this keyword Advanced Search New Results Leg immobilization and subsequent recovery resistance training affect skeletal muscle angiogenesis related markers in young healthy adults regardless of prior resistance training experience Mason C. McIntosh , J. Max Michel , Joshua S. Godwin , Daniel L. Plotkin , Derick A. Anglin , Madison L. Mattingly , Anthony Agyin-Birikorang , Nicholas J. Kontos , Harsimran S. Baweja , View ORCID Profile Matt S. Stock , C. Brooks Mobley , View ORCID Profile Michael D. Roberts doi: https://doi.org/10.1101/2024.11.24.625075 Mason C. McIntosh 1 School of Kinesiology, Auburn University , Auburn, AL USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site J. Max Michel 1 School of Kinesiology, Auburn University , Auburn, AL USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Joshua S. Godwin 1 School of Kinesiology, Auburn University , Auburn, AL USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Daniel L. Plotkin 1 School of Kinesiology, Auburn University , Auburn, AL USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Derick A. Anglin 1 School of Kinesiology, Auburn University , Auburn, AL USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Madison L. Mattingly 1 School of Kinesiology, Auburn University , Auburn, AL USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Anthony Agyin-Birikorang 1 School of Kinesiology, Auburn University , Auburn, AL USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Nicholas J. Kontos 1 School of Kinesiology, Auburn University , Auburn, AL USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Harsimran S. Baweja 1 School of Kinesiology, Auburn University , Auburn, AL USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Matt S. Stock 2 Institute of Exercise Physiology and Rehabilitation Science, School of Kinesiology and Rehabilitation Sciences, University of Central Florida , Orlando, FL, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Matt S. Stock C. Brooks Mobley 1 School of Kinesiology, Auburn University , Auburn, AL USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Michael D. Roberts 1 School of Kinesiology, Auburn University , Auburn, AL USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Michael D. Roberts For correspondence: mdr0024{at}auburn.edu Abstract Full Text Info/History Metrics Preview PDF ABSTRACT We recently reported that resistance trained (T, n=10) and untrained (UT, n=11) young adults experience vastus lateralis (VL) muscle atrophy following two weeks of disuse, and 8 weeks of recovery resistance training (RT) promotes VL hypertrophy in both participant cohorts. However, angiogenesis targets and muscle capillary number were not examined and currently no human studies that have sought to determine if disuse followed by recovery RT affects these outcomes. Thus, we examined whether disuse and/or recovery RT affected these outcomes. All participants underwent two weeks of left leg immobilization using locking leg braces and crutches followed by eight weeks (3d/week) of knee extensor focused progressive RT. VL biopsies were obtained at baseline (PRE), immediately after disuse (MID), and after RT (POST). Western blotting was used to assay angiogenesis markers and immunohistochemistry was performed in 16/21 participants to determine type I and II muscle fiber capillary number. Significant main effects of time (p<0.05) were observed for protein levels of VEGF (MID<POST), VEGFR2 (PRE&MID<POST), TSP-1 (PRE<POST), TIMP1 (MID<POST), phosphorylated/pan eNOS (Ser1177) (POST<PRE), and pan eNOS (PRE<POST). VEGFR2 exhibited a training status*time (p=0.018), but no differences existed between T and UT at any time point. A significant main effect of time was observed for type II fiber capillary number (PRE<POST), and type II fiber cross-sectional area (fCSA) increased from MID to POST (+25%, p<0.001) and PRE to POST (+20%, p=0.019). No significant correlations exist for percentage changes in type II fiber capillary number and type II fCSA from PRE-to-MID (r= 0.020), MID-to-POST (r= 0.392), or PRE-to-POST (r= −0.120) across all participants (p>0.100). Although disuse and recovery RT affect skeletal muscle angiogenesis-related protein targets, prior training history does not differentially affect these outcomes. NEW AND NOTEWORTHY This is the first study to examine how limb immobilization and recovery resistance training affect molecular outcomes related to angiogenesis in younger adults with or without a prior training history. Regardless of resistance training history, the molecular responses are largely similar between participant cohorts and is suggestive of a reduced (pre-mid) and increased (mid-post) angiogenic response, with disuse and subsequent recovery resistance training. INTRODUCTION Skeletal muscle is a tissue that robustly adapts to various loading stimuli ( 1 ), and mechanical overload through resistance training (RT) promotes remodeling to support myofiber and whole tissue hypertrophy ( 2 ). Although the mobilization of satellite cells and anabolic signaling through the mammalian target of rapamycin complex 1 (mTORC1) signaling have been generally appreciated for promoting skeletal muscle hypertrophy ( 3 ), recent evidence indicates angiogenesis (or the process of capillary expansion or growth) may also play a significant role ( 4 ). To this end, longer-term RT promotes increases in skeletal muscle capillarization ( 5 – 7 ), lower pre-training muscle capillary density is associated with limited hypertrophic outcomes in adults ( 8 – 10 ), and genetic mouse models indicate that mechanical overload-induced skeletal muscle hypertrophy is blunted when angiogenesis is impaired ( 11 , 12 ). From a mechanistic standpoint, RT in humans upregulates the mRNA and protein expression of vascular endothelial growth factor (VEGF) ( 13 – 15 ), a potent pro-angiogenic factor ( 4 , 16 ). There is also evidence that RT can dynamically alter matrix metalloproteinases (MMPs) as well as their inhibitors (TIMPs) ( 17 , 18 ), and these proteins are also involved in the extracellular matrix (ECM) remodeling needed for angiogenesis ( 19 ). Finally, limited data indicates that the angiogenesis response may be more robust in untrained (UT) individuals when compared to trained (T) individuals suggesting that diminished angiogenesis may be an adaptive response to RT ( 15 ). Contrary to RT-induced skeletal muscle hypertrophy, disuse promotes remodeling processes that result in skeletal muscle tissue and myofiber atrophy ( 20 ). Disuse related skeletal muscle loss is a common outcome among medically injured patients, those with common illnesses, and aging individuals ( 20 ). Appreciable skeletal muscle atrophy can manifest within 10 days of disuse, equating in the approximate loss of ∼0.5-0.6% of muscle mass per day ( 21 ) and results in up to 30% loss of total quadriceps size after 90-120 days ( 22 , 23 ). These maladaptive responses are, due to altered net protein balance where muscle protein synthesis does not outweigh breakdown, largely due to decreases in muscle protein synthesis ( 24 ). Additional mechanisms associated with disuse-induced skeletal muscle atrophy include a dysregulation in muscle-to-nerve communication, an increase in endoplasmic reticulum stress, and a loss in ribosome content ( 23 – 26 ). Disuse via unilateral leg immobilization has been reported to reduce skeletal muscle capillary content ( 27 ), and this may coincide with the atrophic process. While disuse and mechanical overload potentially operate through different signaling pathways, the molecular response to skeletal muscle disuse and subsequent recovery RT is well-elucidated. Moreover, although RT can increase skeletal muscle capillarization, limited data indicates that the angiogenesis response to RT may be more robust in untrained (UT) individuals when compared to trained (T) individuals ( 15 ), which suggests that diminished angiogenesis may be an adaptive response to RT. Thus, examining the angiogenesis response to disuse and recovery RT in individuals with or without a prior history of resistance training would provide valuable insight. We previously reported that T (n=10) and UT (n=11) young adults experienced similar magnitudes of muscle atrophy according to ultrasound measures following two weeks of disuse, and 8 weeks of recovery RT promoted skeletal muscle hypertrophy in both participant cohorts ( 28 ). However, given that angiogenesis markers were not originally assayed, we sought to determine if the disuse and recovery RT protocol affected several angiogenesis and vascular remodeling markers (VEGF, VEGFR2, TSP-1, MMP2, MMP9, TIMP1, TIMP2, total eNOS, and phosphorylated eNOS) or type I and II muscle fiber capillarization, and whether responses differed between UT and T participants. We developed multiple a priori hypotheses. First, disuse and recovery RT would downregulate and subsequently upregulate skeletal muscle protein levels of VEGF, VEGR2, MMP2, MMP9, and phosphorylated as well as total eNOS (angiogenesis markers) in both UT and T participants, albeit this response would be more robust in UT participants. Second, disuse and recovery RT would upregulate and subsequently downregulate skeletal muscle protein levels of angiogenesis inhibitors (i.e., TSP-1, TIMP1, TIMP2) in both UT and T participants. We additionally hypothesized that changes observed in the protein expression of these angiogenic regulators would be more robust in UT versus T. Finally, we hypothesized muscle capillary number would be greater in T versus UT participants throughout the intervention and that changes in muscle capillary number throughout the intervention would be significantly associated with changes in muscle fiber size. METHODS Ethical approval and participant screening This study is a follow-up investigation of a study approved by the Auburn University Institutional Review Board and in accordance with the most recent revisions of Declaration of Helsinki (IRB protocol #23-220 MR2305, clinical trial registration # NCT05760066 ). The participants were males and females from the local area who met the following criteria: (i) 18-35 years old; (ii) no known cardiometabolic disease (e.g. diabetes, hypertension, heart disease) or any musculoskeletal condition contraindicating participation in exercise training or donating skeletal muscle biopsies; (iii) free from metal implants that would interfere with x-ray based data collection; (iv) had not consumed known anabolic agents that affect hormone status within the past two months (e.g. exogenous testosterone, growth hormone, etc.); (v) free from blood clotting disorders that would contraindicate donating a muscle biopsy; (vi) females were not pregnant or attempting to become pregnant. Additional participant information and protocol for participant recruitment and screening can be found in Michel et al. ( 28 ). Study design A more detailed description of the study design and methodology can be found in Michel et al. ( 28 ). Briefly, UT (n=11, 26±3 years old, 78.4±24.4 kg, 8 males, 3 females) and T (n=10, 27±3 years old, 81.4 ± 8.1 kg, 8 males, 2 females) participants underwent 2 weeks of left leg immobilization via locking brace and crutches followed by 8 weeks of RT. Participants visited the laboratory for baseline data collection and to be fitted for their locking leg brace and crutches (PRE), after two weeks of disuse for subsequent data collection and removal of their locking brace and crutches (MID), and after 8 weeks of RT for final data collection (POST). All data collection sessions occurred a minimum of 72 hours after any RT session and the order of data collection events were held constant for each session. Leg disuse protocol The leg disuse protocol was designed based on previous work by MacLennan et al. ( 29 ). Upon completion of all testing procedures during PRE, participants were fitted with a knee joint immobilizer brace (T Scope® Premier Post-Op Knee Brace; Breg Inc., Carlsbad, CA, USA). The brace was locked in place at ∼90° of knee flexion. In this position the participants’ knee extensors remained relaxed and unloaded to ensure the leg was fully non-weight bearing. Participants were additionally fitted with axillary crutches and were provided gait-training to effectively ambulate with a single leg while using crutches. After two weeks of bracing and ambulation on crutches, each participant returned to the laboratory for Post-brace testing. At this time the brace was removed, and each participant returned to their bilateral full weight bearing gait. Post-brace resistance training protocol Following brace removal, participants underwent 8 weeks of supervised RT at a frequency of three (non-consecutive) days per week. Participants performed barbell back squats, leg extensors, barbell bench press, lat pulldown, and lying hamstring curls on day 1 and 3 of the week. On day 2 of training, the individuals performed leg press, hex-bar deadlift, barbell row, overhead press, and dumbbell biceps curl. Progressive overload was implemented throughout the training period, and additional training details on training can be found in Michel et al. ( 28 ). PRE, MID, and POST testing sessions Urine specific gravity, height, and body mass During testing session visits, participants reported to the laboratory following an overnight fast. Hydration status was assessed via a urine specific gravity test measured with a handheld refractometer (ATAGO; Bellevue, WA, USA) with the goal of ensuring adequate hydration (value ≤1.020) ( 30 ). Body mass and height were assessed with a digital scale (Seca 769, Hanover, MD, USA). Vastus lateralis muscle biopsies After ultrasound scans, skeletal muscle biopsy samples were collected from the left VL a 5 mm Bergstrom needle. Biopsies at MID and POST were taken ∼2 cm proximal from the preceding biopsy scar. Briefly, participants laid supine on an athletic training table and the upper thigh was shaved and cleaned with 70% isopropanol prior to receiving an injection of 1% lidocaine (0.6-0.9 mL). After waiting ∼5 minutes to allow lidocaine to take full effect, the area was cleaned with chlorhexidine and a pilot incision made with a sterile, single-use No. 11 surgical blade (AD Surgical, Sunnyvale, CA, USA). The biopsy was then collected using a 5 mm Bergstrom needle under suction. Approximately 50-100 mg of skeletal muscle tissue was collected, immediately teased of blood and connective tissue, and separated for histological and biochemical analysis. Tissue (∼30 mg) was mounted for histology at a 90° angle on a piece of cork in a 1:1 w/w mixture of optimal cutting temperature (OCT) solution and tragacanth powder (Alfa Aesar, Ward Hill, MA, USA). The mount was then covered in OCT, frozen in liquid nitrogen cooled 2-methylbutane for ∼30 seconds, then contained in a box top floating atop liquid nitrogen before long-term storage at −80°C. A separate ∼20-60 mg tissue sample was placed in foil and flash frozen in liquid nitrogen for protein isolation and western blotting. All tissue triage procedures occurred within a 2-minute window. Wet laboratory analyses Western blotting Muscle tissue (∼20 mg) was lysed using a general cell lysis buffer (Cell Signaling Technology, Danvers, MA, USA; Cat. No. 9803) and tight-fitting pestles. Lysates were then centrifuged at 500 g for 5 minutes and supernatants were placed into new 1.7 mL tubes. A commercially available BCA protein assay kit (Thermo Fisher, Waltham, MA, USA; Cat. No. A55864) and spectrophotometer (Agilent Biotek Synergy H1 hybrid reader; Agilent, Santa Clara, CA, USA) were used to determine supernatant protein concentrations. Thereafter, supernatants were prepared for western blotting at equal protein concentrations (1 µg/µL) using 4x Laemmli buffer and deionized water. Western blot preps (15 µL) were pipetted onto SDS gels (4-15% Criterion TGX Stain-free gels, Bio-Rad Laboratories; Hercules, CA, USA), and proteins were separated by electrophoresis at 180 V for 50 minutes. Proteins were then transferred to methanol-preactivated PVDF membranes (Bio-Rad Laboratories) for 2 hours at 200 mA. Following transfers, membranes were Ponceau stained for 10 minutes, washed with diH 2 O for ∼30 seconds, dried, and digitally imaged (ChemiDoc Touch, Bio-Rad). Following Ponceau imaging, membranes were reactivated in methanol, blocked with 5% non-fat bovine milk in tris-buffered saline with Tween-20 (TBST) for 1 hour, and washed 3x5 minutes in TBST. Membranes were then incubated with primary antibodies (1:1000 dilution in TBST containing 5% bovine serum albumin (BSA)) on a rocker overnight at 4°C (listed in Table 1 ). View this table: View inline View popup Download powerpoint Table 1. Antibodies used for western blotting and immunohistochemistry After overnight primary antibody incubations, antibody solutions were decanted, and membranes were washed for 3x5 minutes in TBST. The membranes were then incubated at room temperature for 60 minutes in in TBST containing 5% BSA and a 1:2000 v/v dilution of HRP-conjugated antibody against the host species of the primary antibody (presented in Table 1 ). The secondary antibody solution was decanted, and membranes were washed for 3x5 minutes in TBST. The membranes were then developed in a gel documentation system (ChemiDoc Touch, Bio-Rad) with enhanced chemiluminescent reagent (Luminata Forte HRP substrate; Millipore Sigma, Burlington, MA, USA), and band densitometry was performed using associated software. For non-phosphorylated targets, target band densities were obtained and divided by Ponceau densitometry values and fold-change values were derived by dividing Ponceau-normalized band density values by the aggregate PRE mean value of the UT group for each target. Band density values for phosphorylated eNOS was divided by corresponding pan band density values (after being normalized to ponceau stains), and again fold-change values were derived by dividing these values by the aggregate PRE mean value of the of T participants. Immunohistochemistry for capillarization quantification Biopsy samples preserved with optimal cutting temperature media were sectioned at a thickness of 12 μm using a cryotome (Leica Biosystems; Buffalo Grove, IL, USA), adhered to positively charged glass slides (VWR; Radnor, PA, USA), and stored at −80℃ until being batch processed for immunohistochemical analyses to detect capillaries, fiber type, dystrophin, and nuclei. During batch-processing, sections were removed from −80°C storage, air-dried at room temperature for ≥2 hours, and fixed with acetone at −20°C for 5 minutes. Slides were then incubated with 3% hydrogen peroxide for 10 minutes at room temperature, followed by a 1-minute incubation with autofluorescence quenching reagent (TrueBlack, Cat. No. 23007; Biotium, Fremont, CA, USA) and blocked for 1 hour in 2.5% horse serum in phosphate-buffered saline (PBS) at room temperature. After blocking, slides were incubated overnight at 4°C with a primary antibody cocktail in PBS containing 2.5% horse serum and 1:100 v/v dilutions of anti-PECAM-1, anti-type I myosin heavy chain, and anti-dystrophin antibodies listed in Table 1 . The following day, the sections were washed with PBS 3x5 minutes and incubated for 60 minutes in a secondary antibody cocktail containing 1:250 v/v dilutions of the fluorophore-conjugated secondary antibodies listed in Table 1 . Slides were then stained with DAPI (1:10,000; 4’, 6-diamidino-2-phenylindole; Cat. No. D3571; Thermo Fisher Scientific) for 10 minutes at room temperature and mounted with glass coverslips using 1:1 PBS and glycerol as mounting medium. Sections were stored in the dark at 4°C until imaging was conducted. Multiple digital 20x images per participants’ time points were captured with a fluorescence microscope at (Zeiss Axio Imager.M2) and motorized stage. All areas selected for analyses were free of freeze-fracture artifacts. Fiber cross-sectional area was conducted in Michel et al. ( 28 ). Capillary contacts to type I or type II fibers were manually quantified using a tally counter. Statistical analyses Data were plotted and analyzed in GraphPad Prism (v10.2.2). Data were first checked for normality using Shapiro-Wilk tests. Only VEGF protein in T participants, MMP9 in UT, and TIMP1 in UT were non-normally distributed (p<0.05). Thus, we opted to proceed with parametric analyses given that most dependent variables were normally distributed. All outcome variables were examined using two-way (training status*time) repeated measures ANOVAs with, and Tukey’s post hoc tests used to decompose significant main effects and/or interactions. Select Pearson’s correlations were conducted on certain change scores from immunohistochemistry analysis as explained in the results section. Statistical significance was established throughout as p <0.05. RESULTS Angiogenesis markers Figure 1 contains skeletal muscle protein levels of positive (VEGF, VEGFR2) and negative (TSP-1) regulators of angiogenesis. VEGF exhibited a main effect of time (p=0.006; Fig. 1a ) where PRE and POST values were greater than MID (p < 0.011; Fig. 1a ). VEGFR2 exhibited a main effect of time (p<0.001) and a training status*time interaction (p=0.018; Fig. 1b ). Although PRE values of the trained participants trended higher than the untrained participants (p=0.083), values between training status groups were not significantly different at MID (p=0.694) or POST (p=0.127). Values at PRE and MID, however, were less than POST in both training status groups (p < 0.047). TSP-1 exhibited a main effect of time (p=0.040) and a training status*time interaction (p=0.038; Fig. 1c ). Despite a significant interaction, values were not different between T and UT at any timepoint (p≥.0.079), while values at PRE were less than MID in both training status groups (p < 0.026). Download figure Open in new tab Figure 1. Levels of proteins associated with angiogenesis Data presented as mean ± standard deviation for VEGF (a), VEGFR2 (b), and TSP-1 (c) prior to the intervention (Pre), following two weeks of leg disuse (Mid), and following 8 weeks of recovery resistance training (Post). Panel d shows representative western blots. Abbreviations: T, trained participant; UT, untrained participant. Other note: each participant’s data point is overlain on bar graphs and mean values are depicted at bottom of bar. Extracellular matrix remodeling markers Figure 2 contains skeletal muscle protein expression data from select matrix metalloproteinases (MMPs) and MMP inhibitors (TIMP1/2) implicated in angiogenesis. Neither MMP2 ( Fig. 2a ), MMP9 ( Fig. 2b ), TIMP1 ( Fig. 2c ), nor TIMP2 ( Fig. 2d ) exhibited significant main or interaction effects. Download figure Open in new tab Figure 2. Levels of proteins associated with extracellular matrix remodeling during angiogenesis Data presented as mean ± standard deviation for MMP2 (a), MMP9 (b), TIMP1 (c) and TIMP2 prior to the intervention (Pre), following two weeks of leg disuse (Mid), and following 8 weeks of recovery resistance training (Post). Panel e shows representative western blots. Abbreviations: T, trained participant; UT, untrained participant. Other note: each participant’s data point is overlain on bar graphs and mean values are depicted at bottom of bar. eNOS markers Figure 3 contains skeletal muscle pan and phosphorylated eNOS levels. While significant interaction effects were not evident for these markers, significant main effects of time were evident for both. Specifically, phosphorylated eNOS levels were lower in all participants at POST versus PRE (p=0.017; Fig. 3a ) whereas pan eNOS levels were greater at POST versus PRE (p=0.044) and MID (p=0.001; Fig. 3b ). Download figure Open in new tab Figure 3. Phosphorylated and pan eNOS responses Data presented as mean ± standard deviation for phosphorylated-eNOS (a) and pan protein levels (b) prior to the intervention (Pre), following two weeks of leg disuse (Mid), and following 8 weeks of recovery resistance training (Post). Panel c shows representative western blots. Abbreviations: T, trained participant; UT, untrained participant. Other note: each participant’s data point is overlain on bar graphs and mean values are depicted at bottom of bar. Changes in capillaries per fiber Figure 4 contains skeletal muscle type I and II myofiber capillary count data. Type I capillary number did not exhibit significant main or interaction effects ( Fig. 4a ). However, Type II capillary number did exhibit a significant main effect of time whereby POST was greater than PRE in all participants (p=0.014, Fig. 4b ). Download figure Open in new tab Figure 4. Type I and II myofiber capillary responses Data presented as mean ± standard deviation for type I (a) and type II (b) myofiber capillary number prior to the intervention (Pre), following two weeks of leg disuse (Mid), and following 8 weeks of recovery resistance training (Post). Panel c shows a representative 20x fluorescent image of PECAM-1 only and a merged image; note, dystrophin (Cy5) is pseudocolored white. Abbreviations: T, trained participant; UT, untrained participant. Other note: each participant’s data point is overlain on bar graphs and mean values are depicted at bottom of bar. Associations between changes in type II fiber capillary number and myofiber size Given that type II fiber capillary number increased with RT regardless of training group, we were interested in determining if this outcome was associated with percentage changes in type II fiber cross-sectional area (fCSA) values. Michel et al. reported type II fCSA values in 21 participants, and the 16 participants assayed herein demonstrated numerical decreases from PRE to MID (6225±1593 to 5943±1566 µm 2 , p=0.137) as well as values at POST (7165±1418 µm 2 ) that were greater than PRE (p=0.019) and MID (p<0.001). However, no significant correlations existed for percentage changes in type II fiber capillary number and type II fCSA from PRE-to-MID (r= 0.020, p=0.941), MID-to-POST (r= 0.392, p=0.133), or PRE-to-POST (r= −0.120, p=0.657). DISCUSSION Reduced skeletal muscle angiogenesis and losses in capillary content (i.e., capillary rarefaction) have been posited to play a role in skeletal muscle disuse-induced atrophy ( 31 , 32 ). Moreover, various skeletal muscle atrophy models in rodents suggest capillary rarefaction and/or suppressed angiogenesis coincide with skeletal muscle atrophy ( 25 , 33 – 38 ). Various rodent and human studies have sought to assess how reloading following disuse atrophy effects various markers related to angiogenesis ( 39 – 42 ). This is highlighted in that evidence in human participants suggests that 4-6 weeks of endurance training following two weeks of leg immobilization alters markers suggestive of increased angiogenesis ( 27 , 43 ). However, no human study to date has examined how recovery RT following disuse atrophy affects these outcomes. Therefore, this study aimed to assess alterations in markers associated with skeletal muscle angiogenesis following two weeks of leg immobilization immediately followed by 8 weeks of subsequent recovery RT. In agreement with our hypotheses, 2 weeks of leg immobilization is seemingly anti-angiogenic as evidenced through a decrease in VEGF and increase in TSP-1 protein levels. Also, in agreement with our hypotheses, 8 weeks of recovery RT enhances angiogenesis as evidenced through MID-to-POST increases in VEGF and VEGFR2 protein levels as well as a PRE-to-POST increase in type II fiber capillary number. Prior to a more expanded discussion of the implications of our findings, it is worth noting that training history differences in most outcomes were not evident. This finding also agrees with the parent publication by Michel et al. ( 28 ) who reported that atrophic and hypertrophic responses, along with corresponding molecular targets, were similar regardless of training status/history in the given model. Extracellular matrix remodeling markers associated with angiogenesis remained relatively unchanged with disuse and subsequent recovery RT, albeit TIMP1 protein levels were responsive across time points. Several factors directly related to angiogenesis (VEGF, VEGFR2, and TSP-1) were dynamically altered following 2 weeks of disuse. TSP-1 is a potent inhibitor of angiogenesis by antagonizing VEGF and reducing endothelial cell proliferation and migration ( 19 , 44 ). In rodent models, skeletal muscle capillary number is reduced with the administration of a TSP-1 mimetic ( 45 ), and conversely, capillary number is elevated in a global TSP-1 knockout model ( 46 ). Interestingly, our findings that two weeks of leg immobilization reduce VEGF and increase TSP-1 protein levels in skeletal muscle agree with another human study demonstrating similar effects following two weeks of leg bracing ( 43 ). However, in the current model of disuse, we did not observe capillary rarefaction despite decreases in skeletal muscle VEGF and VEGFR2 as well as an increase in TSP-1. Our VEGF, VEGFR2 and TSP-1 data suggest that observed protein expression patterns precede marked changes in ( 19 ) and that skeletal muscle capillary rarefaction may be observed with a longer duration or with a more severe disuse model/stimulus. Resistance training consistently leads to skeletal muscle hypertrophy, and this has been shown to coincide with increased skeletal muscle capillarization ( 5 , 7 , 47 – 52 ) along with acute upregulations of angiogenic markers after a single bout or short-term RT ( 5 , 13 – 15 , 53 ), Furthermore, eNOS (an enzyme known to promote vasodilation and angiogenesis) is also altered with exercise ( 16 , 19 ). In the current study, we report an increase in VEGF, VEGFR2, and eNOS protein levels from MID to POST time points. This suggests dynamic interplay between angiogenic markers and following disuse atrophy and subsequent recovery RT. The observed increases in type II myofiber capillary number agree with other human studies reporting that RT increases skeletal muscle capillary number ( 5 , 48 , 49 , 54 ). Despite increases in capillary number, no associations were shown between percentage changes in fCSA and capillary number in the current study. In contrast with our data, skeletal muscle capillarization has been shown to be predictive of the skeletal muscle hypertrophic response to RT among older adults ( 8 , 9 ). While speculative, our lack of associations may be due to younger/healthy adults being investigated compared to the studies previously mentioned. In the current model, several markers known to regulate angiogenesis are dynamically altered following both disuse and recovery RT, while protein levels of these markers are affected, capillary number is only significantly altered in type II myofibers following RT and is not associated with myofiber hypertrophy. Taken together, the data suggest that angiogenic-related markers are altered in both an atrophic and hypertrophy stimulus but do not seem to drive changes in skeletal muscle mass in a young healthy population. Further interrogations comparing different aging populations following both disuse and RT stimulus are needed to fully unveil any further associations between skeletal muscle capillarization and skeletal muscle hypertrophic adaptations. Limitations Similar to many human participant RT studies, this study is limited by a small sample size. A larger sample size with a more diverse participant pool would likely increase the generalizability of our findings. Moreover, the participants herein were predominantly males who underwent non-complicated disuse whereby the braced limb was not injured, and the immobilization period was among a less severe model of immobilization (e.g., bedrest, limb casting, microgravity). Finally, biopsies were only performed in the basal state, so we did not interrogate markers of gene regulation (e.g., mRNA expression) related to angiogenesis that might be acutely responsive to RT and/or disuse atrophy. Indeed, these markers would add mechanistic insight to how the model may affect phenotypic outcomes. Conclusions This study furthers findings from our original study by Michel et al. ( 28 ) by providing the first human evidence demonstrating that leg immobilization and subsequent recovery RT dynamically alter skeletal muscle markers associated with angiogenesis in younger adults. However, further investigation is needed to determine if our findings are consistent across other disuse models (e.g., complicated disuse with casting or bedrest) and different participant cohorts (e.g., older individuals). ADDITIONAL INFORMATION CONFLICTS OF INTEREST The authors declare they have no competing interests in relation to these data. FUNDING STATEMENT D.L.P. was supported by an Auburn University Presidential Research Fellowship, D.A.A. was supported by an Auburn University Presidential Opportunity Fellowship, and M.C.M. was supported by a NIH fellowship (5T32GM141739-03). Funding for this project was provided by discretionary laboratory funds from MDR. DATA AVAILABILITY STATEMENT Data that support the findings of this study are available from the corresponding author ( mdr0024{at}auburn.edu ) upon reasonable request. CLINICAL TRIAL REGISTRATION This study was registered as a clinical trial at clinicaltrials.gov ( NCT05760066 ). ETHICS APPROVAL STATEMENT Study protocols were carried out in accordance with the most recent version of the declaration of Helsinki. All study procedures were approved by Auburn University’s Institutional Review Board (approval number: 23-220 MR 2305). PATIENT CONSENT STATEMENT All participants in this study provided verbal and written consent in accordance with the above IRB approval. AUTHOR CONTRIBUTIONS MCM and MDR primarily drafted the manuscript and prepared figures. HSB is a licensed physical therapist who assisted with oversight throughout the intervention. MSS provided critical equipment and expertise for the carrying out of the leg immobilization protocol. MCM primarily carried out laboratory-based assays. JMM, JSG, DLP, DAA, MLM, AAB, NJK, and CBM provided crucial assistance with original data collection procedures and/or consistent input for the duration of this follow-up investigation. All co-authors assisted with revising and editing the manuscript, and all co-authors approved the final version. ACKNOWLEDGEMENTS We would like to acknowledge the participants who completed this study. REFERENCES 1. ↵ Joanisse S , Lim C , McKendry J , McLeod JC , Stokes T , and Phillips SM . Recent advances in understanding resistance exercise training-induced skeletal muscle hypertrophy in humans . F1000Res 9 : 2020 . 2. ↵ Roberts MD , McCarthy JJ , Hornberger TA , Phillips SM , Mackey AL , Nader GA , Boppart MD , Kavazis AN , Reidy PT , Ogasawara R , Libardi CA , Ugrinowitsch C , Booth FW , and Esser KA . Mechanisms of mechanical overload-induced skeletal muscle hypertrophy: current understanding and future directions . Physiol Rev 103 : 2679 – 2757 , 2023 . OpenUrl CrossRef PubMed 3. ↵ McIntosh MC , Sexton CL , Godwin JS , Ruple BA , Michel JM , Plotkin DL , Ziegenfuss TN , Lopez HL , Smith R , Dwaraka VB , Sharples AP , Dalbo VJ , Mobley CB , Vann CG , and Roberts MD . Different Resistance Exercise Loading Paradigms Similarly Affect Skeletal Muscle Gene Expression Patterns of Myostatin-Related Targets and mTORC1 Signaling Markers . Cells 12 : 2023 . 4. ↵ McIntosh MC , Anglin DA , Robinson AT , Beck DT , and Roberts MD . Making the case for resistance training in improving vascular function and skeletal muscle capillarization . Front Physiol 15 : 1338507 , 2024 . OpenUrl CrossRef PubMed 5. ↵ Holloway TM , Snijders T , J VANK , LJC Vanl , and Verdijk LB . Temporal Response of Angiogenesis and Hypertrophy to Resistance Training in Young Men . Med Sci Sports Exerc 50 : 36 – 45 , 2018 . OpenUrl 6. Campos GE , Luecke TJ , Wendeln HK , Toma K , Hagerman FC , Murray TF , Ragg KE , Ratamess NA , Kraemer WJ , and Staron RS . Muscular adaptations in response to three different resistance-training regimens: specificity of repetition maximum training zones . Eur J Appl Physiol 88 : 50 – 60 , 2002 . OpenUrl CrossRef PubMed Web of Science 7. ↵ McCall GE , Byrnes WC , Dickinson A , Pattany PM , and Fleck SJ . Muscle fiber hypertrophy, hyperplasia, and capillary density in college men after resistance training . J Appl Physiol (1985) 81 : 2004 – 2012 , 1996 . OpenUrl CrossRef PubMed Web of Science 8. ↵ Moro T , Brightwell CR , Phalen DE , McKenna CF , Lane SJ , Porter C , Volpi E , Rasmussen BB , and Fry CS . Low skeletal muscle capillarization limits muscle adaptation to resistance exercise training in older adults . Exp Gerontol 127 : 110723 , 2019 . 9. ↵ Snijders T , Nederveen JP , Joanisse S , Leenders M , Verdijk LB , van Loon LJ , and Parise G . Muscle fibre capillarization is a critical factor in muscle fibre hypertrophy during resistance exercise training in older men . J Cachexia Sarcopenia Muscle 8 : 267 – 276 , 2017 . OpenUrl CrossRef PubMed 10. ↵ Thomas ACQ , Brown A , Hatt AA , Manta K , Costa-Parke A , Kamal M , Joanisse S , McGlory C , Phillips SM , Kumbhare D , and Parise G . Short-term aerobic conditioning prior to resistance training augments muscle hypertrophy and satellite cell content in healthy young men and women . FASEB J 36 : e22500 , 2022 . OpenUrl CrossRef PubMed 11. ↵ Ato S , Fukada SI , Kokubo H , and Ogasawara R . Implication of satellite cell behaviors in capillary growth via VEGF expression-independent mechanism in response to mechanical loading in HeyL-null mice . Am J Physiol Cell Physiol 322 : C275 – C282 , 2022 . OpenUrl CrossRef PubMed 12. ↵ Huey KA , Smith SA , Sulaeman A , and Breen EC . Skeletal myofiber VEGF is necessary for myogenic and contractile adaptations to functional overload of the plantaris in adult mice . J Appl Physiol (1985) 120 : 188 – 195 , 2016 . OpenUrl CrossRef PubMed 13. ↵ Gavin TP , Drew JL , Kubik CJ , Pofahl WE , and Hickner RC . Acute resistance exercise increases skeletal muscle angiogenic growth factor expression . Acta Physiol (Oxf ) 191 : 139 – 146 , 2007 . OpenUrl CrossRef PubMed Web of Science 14. Jensen L , Pilegaard H , Neufer PD , and Hellsten Y . Effect of acute exercise and exercise training on VEGF splice variants in human skeletal muscle . Am J Physiol Regul Integr Comp Physiol 287 : R397 – 402 , 2004 . OpenUrl CrossRef PubMed Web of Science 15. ↵ Richardson RS , Wagner H , Mudaliar SR , Saucedo E , Henry R , and Wagner PD . Exercise adaptation attenuates VEGF gene expression in human skeletal muscle . Am J Physiol Heart Circ Physiol 279 : H772 – 778 , 2000 . OpenUrl CrossRef PubMed Web of Science 16. ↵ Olfert IM , Baum O , Hellsten Y , and Egginton S . Advances and challenges in skeletal muscle angiogenesis . Am J Physiol Heart Circ Physiol 310 : H326 – 336 , 2016 . OpenUrl CrossRef PubMed 17. ↵ Scarpelli MC , Bergamasco JGA , Godwin JS , Mesquita PHC , Chaves TS , Silva DG , Bittencourt D , Dias NF , Medalha Junior RA , Carello Filho PC , Angleri V , Costa LAR , Kavazis AN , Ugrinowitsch C , Roberts MD , and Libardi CA . Correction to: Resistance training-induced changes in muscle proteolysis and extracellular matrix remodeling biomarkers in the untrained and trained states . Eur J Appl Physiol 124 : 2763 , 2024 . OpenUrl CrossRef PubMed 18. ↵ Godwin JS , Sexton CL , Kontos NJ , Ruple BA , Willoughby DS , Young KC , Mobley CB , and Roberts MD . Extracellular matrix content and remodeling markers do not differ in college-aged men classified as higher and lower responders to resistance training . J Appl Physiol (1985) 134 : 731 – 741 , 2023 . OpenUrl CrossRef PubMed 19. ↵ Ross M , Kargl CK , Ferguson R , Gavin TP , and Hellsten Y . Exercise-induced skeletal muscle angiogenesis: impact of age, sex, angiocrines and cellular mediators . Eur J Appl Physiol 123 : 1415 – 1432 , 2023 . OpenUrl CrossRef PubMed 20. ↵ Nunes EA , Stokes T , McKendry J , Currier BS , and Phillips SM . Disuse-induced skeletal muscle atrophy in disease and nondisease states in humans: mechanisms, prevention, and recovery strategies . Am J Physiol Cell Physiol 322 : C1068 – C1084 , 2022 . OpenUrl CrossRef PubMed 21. ↵ Wall BT , Dirks ML , and van Loon LJ . Skeletal muscle atrophy during short-term disuse: implications for age-related sarcopenia . Ageing Res Rev 12 : 898 – 906 , 2013 . OpenUrl CrossRef PubMed 22. ↵ Preobrazenski N , Seigel J , Halliday S , Janssen I , and McGlory C . Single-leg disuse decreases skeletal muscle strength, size, and power in uninjured adults: A systematic review and meta-analysis . J Cachexia Sarcopenia Muscle 14 : 684 – 696 , 2023 . OpenUrl CrossRef PubMed 23. ↵ Sirago G , Pellegrino MA , Bottinelli R , Franchi MV , and Narici MV . Loss of neuromuscular junction integrity and muscle atrophy in skeletal muscle disuse . Ageing Res Rev 83 : 101810 , 2023 . 24. ↵ Michel JM , Hettinger Z , Ambrosio F , Egan B , Roberts MD , Ferrando AA , Graham ZA , and Bamman MM . Mitigating skeletal muscle wasting in unloading and augmenting subsequent recovery . J Physiol 2024 . 25. ↵ He Z , Song Q , Yu Y , Liu F , Zhao J , Un W , Da X , Xu C , Yao Y , and Wang QK . Protein therapy of skeletal muscle atrophy and mechanism by angiogenic factor AGGF1 . J Cachexia Sarcopenia Muscle 14 : 978 – 991 , 2023 . OpenUrl CrossRef PubMed 26. ↵ Sharlo K , Tyganov SA , Tomilovskaya E , Popov DV , Saveko AA , and Shenkman BS . Effects of Various Muscle Disuse States and Countermeasures on Muscle Molecular Signaling . Int J Mol Sci 23 : 2021 . 27. ↵ Vigelso A , Gram M , Wiuff C , Andersen JL , Helge JW , and Dela F . Six weeks’ aerobic retraining after two weeks’ immobilization restores leg lean mass and aerobic capacity but does not fully rehabilitate leg strength in young and older men . J Rehabil Med 47 : 552 – 560 , 2015 . OpenUrl CrossRef PubMed 28. ↵ Michel JM , Godwin JS , Plotkin DL , McIntosh MC , Mattingly ML , Agostinelli PJ , Mueller BJ , Anglin DA , Berry AC , and Vega MM . Effects of leg immobilization and recovery resistance training on skeletal muscle-molecular markers in previously resistance trained versus untrained adults . bioRxiv 2024 . 29. ↵ MacLennan RJ , Sahebi M , Becker N , Davis E , Garcia JM , and Stock MS . Declines in skeletal muscle quality vs. size following two weeks of knee joint immobilization . PeerJ 8 : e8224 , 2020 . OpenUrl CrossRef PubMed 30. ↵ McDermott BP , Anderson SA , Armstrong LE , Casa DJ , Cheuvront SN , Cooper L , Kenney WL , O’Connor FG , and Roberts WO . National Athletic Trainers’ Association Position Statement: Fluid Replacement for the Physically Active . J Athl Train 52 : 877 – 895 , 2017 . OpenUrl CrossRef PubMed 31. ↵ Hendrickse PW , Wust RCI , Ganse B , Giakoumaki I , Rittweger J , Bosutti A , and Degens H . Capillary rarefaction during bed rest is proportionally less than fibre atrophy and loss of oxidative capacity . J Cachexia Sarcopenia Muscle 13 : 2712 – 2723 , 2022 . OpenUrl CrossRef PubMed 32. ↵ Brooks NE , and Myburgh KH . Skeletal muscle wasting with disuse atrophy is multi-dimensional: the response and interaction of myonuclei, satellite cells and signaling pathways . Front Physiol 5 : 99 , 2014 . 33. ↵ Oguri G , Ikegami R , Ugawa H , Katoh M , Obi S , Sakuma M , Takeda N , Kano Y , Toyoda S , and Nakajima T . Muscle Atrophy and mRNA-miRNA Network Analysis of Vascular Endothelial Growth Factor (VEGF) in a Mouse Model of Denervation-Induced Disuse . Cureus 16 : e68974 , 2024 . OpenUrl 34. Tanaka M , Kanazashi M , Kondo H , and Fujino H . Time course of capillary regression and an expression balance between vascular endothelial growth factor-A and thrombospondin-1 in the soleus muscle of hindlimb unloaded rats . Muscle Nerve 65 : 350 – 360 , 2022 . OpenUrl CrossRef PubMed 35. Paudyal A , Slevin M , Maas H , and Degens H . Time course of denervation-induced changes in gastrocnemius muscles of adult and old rats . Exp Gerontol 106 : 165 – 172 , 2018 . OpenUrl CrossRef PubMed 36. Nakagawa K , Tamaki H , Hayao K , Yotani K , Ogita F , Yamamoto N , and Onishi H . Electrical Stimulation of Denervated Rat Skeletal Muscle Retards Capillary and Muscle Loss in Early Stages of Disuse Atrophy . Biomed Res Int 2017: 5695217 , 2017 . 37. Kanazashi M , Okumura Y , Al-Nassan S , Murakami S , Kondo H , Nagatomo F , Fujita N , Ishihara A , Roy RR , and Fujino H . Protective effects of astaxanthin on capillary regression in atrophied soleus muscle of rats . Acta Physiol (Oxf ) 207 : 405 – 415 , 2013 . OpenUrl CrossRef PubMed 38. ↵ Roudier E , Gineste C , Wazna A , Dehghan K , Desplanches D , and Birot O . Angio-adaptation in unloaded skeletal muscle: new insights into an early and muscle type-specific dynamic process . J Physiol 588 : 4579 – 4591 , 2010 . OpenUrl CrossRef PubMed 39. ↵ Oliveira JRS , Mohamed JS , Myers MJ , Brooks MJ , and Alway SE . Effects of hindlimb suspension and reloading on gastrocnemius and soleus muscle mass and function in geriatric mice . Exp Gerontol 115 : 19 – 31 , 2019 . OpenUrl CrossRef PubMed 40. Chacon-Cabrera A , Lund-Palau H , Gea J , and Barreiro E . Time-Course of Muscle Mass Loss, Damage, and Proteolysis in Gastrocnemius following Unloading and Reloading: Implications in Chronic Diseases . PLoS One 11 : e0164951 , 2016 . OpenUrl CrossRef PubMed 41. Chacon-Cabrera A , Gea J , and Barreiro E . Short- and Long-Term Hindlimb Immobilization and Reloading: Profile of Epigenetic Events in Gastrocnemius . J Cell Physiol 232 : 1415 – 1427 , 2017 . OpenUrl CrossRef PubMed 42. ↵ Heinemeier KM , Olesen JL , Haddad F , Schjerling P , Baldwin KM , and Kjaer M . Effect of unloading followed by reloading on expression of collagen and related growth factors in rat tendon and muscle . J Appl Physiol (1985) 106 : 178 – 186 , 2009 . OpenUrl CrossRef PubMed Web of Science 43. ↵ Gliemann L , Rytter N , Jorgensen TS , Piil P , Carter H , Nyberg M , Grassi M , Daumer M , and Hellsten Y . The Impact of Lower Limb Immobilization and Rehabilitation on Angiogenic Proteins and Capillarization in Skeletal Muscle . Med Sci Sports Exerc 53 : 1797 – 1806 , 2021 . OpenUrl CrossRef 44. ↵ Lawler PR , and Lawler J . Molecular basis for the regulation of angiogenesis by thrombospondin-1 and -2 . Cold Spring Harb Perspect Med 2 : a006627 , 2012 . 45. ↵ Audet GN , Fulks D , Stricker JC , and Olfert IM . Chronic delivery of a thrombospondin-1 mimetic decreases skeletal muscle capillarity in mice . PLoS One 8 : e55953 , 2013 . OpenUrl CrossRef PubMed 46. ↵ Malek MH , and Olfert IM . Global deletion of thrombospondin-1 increases cardiac and skeletal muscle capillarity and exercise capacity in mice . Exp Physiol 94 : 749 – 760 , 2009 . OpenUrl CrossRef PubMed Web of Science 47. ↵ Holloway TM , Morton RW , Oikawa SY , McKellar S , Baker SK , and Phillips SM . Microvascular adaptations to resistance training are independent of load in resistance-trained young men . Am J Physiol Regul Integr Comp Physiol 315 : R267 – R273 , 2018 . OpenUrl CrossRef PubMed 48. ↵ Nederveen JP , Snijders T , Joanisse S , Wavell CG , Mitchell CJ , Johnston LM , Baker SK , Phillips SM , and Parise G . Altered muscle satellite cell activation following 16 wk of resistance training in young men . Am J Physiol Regul Integr Comp Physiol 312 : R85 – R92 , 2017 . OpenUrl CrossRef PubMed 49. ↵ Verdijk LB , Snijders T , Holloway TM , J VANK , and LJ Vanl . Resistance Training Increases Skeletal Muscle Capillarization in Healthy Older Men . Med Sci Sports Exerc 48 : 2157 – 2164 , 2016 . OpenUrl CrossRef 50. Jensen L , Bangsbo J , and Hellsten Y . Effect of high intensity training on capillarization and presence of angiogenic factors in human skeletal muscle . J Physiol 557 : 571 – 582 , 2004 . OpenUrl CrossRef PubMed Web of Science 51. Hostler D , Schwirian CI , Campos G , Toma K , Crill MT , Hagerman GR , Hagerman FC , and Staron RS . Skeletal muscle adaptations in elastic resistance-trained young men and women . Eur J Appl Physiol 86 : 112 – 118 , 2001 . OpenUrl CrossRef PubMed 52. ↵ Green H , Goreham C , Ouyang J , Ball-Burnett M , and Ranney D . Regulation of fiber size, oxidative potential, and capillarization in human muscle by resistance exercise . Am J Physiol 276 : R591 – 596 , 1999 . OpenUrl CrossRef 53. ↵ Gustafsson T , Puntschart A , Kaijser L , Jansson E , and Sundberg CJ . Exercise-induced expression of angiogenesis-related transcription and growth factors in human skeletal muscle . Am J Physiol 276 : H679 – 685 , 1999 . OpenUrl PubMed Web of Science 54. ↵ Cocks M , Shaw CS , Shepherd SO , Fisher JP , Ranasinghe AM , Barker TA , Tipton KD , and Wagenmakers AJ . Effect of resistance training on microvascular density and eNOS content in skeletal muscle of sedentary men . Microcirculation 21 : 738 – 746 , 2014 . OpenUrl CrossRef PubMed View the discussion thread. Back to top Previous Next Posted November 26, 2024. Download PDF Email Thank you for your interest in spreading the word about bioRxiv. NOTE: Your email address is requested solely to identify you as the sender of this article. Your Email * Your Name * Send To * Enter multiple addresses on separate lines or separate them with commas. You are going to email the following Leg immobilization and subsequent recovery resistance training affect skeletal muscle angiogenesis related markers in young healthy adults regardless of prior resistance training experience Message Subject (Your Name) has forwarded a page to you from bioRxiv Message Body (Your Name) thought you would like to see this page from the bioRxiv website. Your Personal Message CAPTCHA This question is for testing whether or not you are a human visitor and to prevent automated spam submissions. Share Leg immobilization and subsequent recovery resistance training affect skeletal muscle angiogenesis related markers in young healthy adults regardless of prior resistance training experience Mason C. McIntosh , J. Max Michel , Joshua S. Godwin , Daniel L. Plotkin , Derick A. Anglin , Madison L. Mattingly , Anthony Agyin-Birikorang , Nicholas J. Kontos , Harsimran S. Baweja , Matt S. Stock , C. Brooks Mobley , Michael D. Roberts bioRxiv 2024.11.24.625075; doi: https://doi.org/10.1101/2024.11.24.625075 Share This Article: Copy Citation Tools Leg immobilization and subsequent recovery resistance training affect skeletal muscle angiogenesis related markers in young healthy adults regardless of prior resistance training experience Mason C. McIntosh , J. Max Michel , Joshua S. Godwin , Daniel L. Plotkin , Derick A. Anglin , Madison L. Mattingly , Anthony Agyin-Birikorang , Nicholas J. Kontos , Harsimran S. Baweja , Matt S. Stock , C. Brooks Mobley , Michael D. Roberts bioRxiv 2024.11.24.625075; doi: https://doi.org/10.1101/2024.11.24.625075 Citation Manager Formats BibTeX Bookends EasyBib EndNote (tagged) EndNote 8 (xml) Medlars Mendeley Papers RefWorks Tagged Ref Manager RIS Zotero Tweet Widget Facebook Like Google Plus One Subject Area Physiology Subject Areas All Articles Animal Behavior and Cognition (7651) Biochemistry (17746) Bioengineering (13928) Bioinformatics (42066) Biophysics (21499) Cancer Biology (18650) Cell Biology (25579) Clinical Trials (138) Developmental Biology (13409) Ecology (19947) Epidemiology (2067) Evolutionary Biology (24374) Genetics (15633) Genomics (22557) Immunology (17775) Microbiology (40505) Molecular Biology (17217) Neuroscience (88796) Paleontology (667) Pathology (2845) Pharmacology and Toxicology (4836) Physiology (7664) Plant Biology (15179) Scientific Communication and Education (2047) Synthetic Biology (4304) Systems Biology (9839) Zoology (2272)