8-week Time Point Research Articles

Brief Peer-Supported Web-Based Skills Training in Affective and Interpersonal Regulation (BPS webSTAIR) for Trauma-Exposed Veterans in the Community: Randomized Controlled Trial.

Peer-supported mobile health (mHealth) programs hold the promise of providing a low-burden approach to increasing access to care and improving mental health. While peer support has been shown to improve engagement in care, there is limited investigation into the impact of peers on symptom outcomes. Trauma-exposed populations frequently endure co-occurring posttraumatic stress and depressive symptoms as well as difficulties in day-to-day functioning. This study evaluated the potential benefits of a peer-supported, transdiagnostic mHealth program on symptom outcomes and functioning. This randomized controlled trial tested the effectiveness of Brief Peer-Supported (BPS) web-based Skills Training in Affective and Interpersonal Regulation (webSTAIR), a 6-module transdiagnostic digital program derived from Skills Training in Affective and Interpersonal Regulation and compared to waitlist control in a community sample of veterans who screened positive for either posttraumatic stress disorder (PTSD) or depression. A total of 178 veterans were enrolled in this study using a 2:1 randomization scheme with 117 assigned to BPS webSTAIR and 61 assigned to waitlist control. PTSD and depressive symptoms as well as emotion regulation and psychosocial functioning were assessed at pretreatment, posttreatment, and 8-week follow-up time points. Mixed-effects models were used to assess change in outcome measures across time points. Exploratory analyses were conducted to determine whether the type and number of peer interactions influenced outcomes. Significant interaction effects were observed for all outcomes such that participants randomized to BPS webSTAIR reported significantly greater improvement at the posttreatment time point compared to waitlist control with moderate effect sizes for PTSD (d=0.48), depression (d=0.64), emotion regulation (d=0.61), and functional impairment (d=0.61); gains were maintained at 8-week follow-up. An initial cohort of participants who were required to engage with a peer coach to progress through the modules interacted more frequently with peers but completed fewer modules compared to a later cohort for whom peer engagement was optional. Overall, those who completed more modules reported greater improvement in all outcomes. BPS webSTAIR was effective in improving PTSD and depression symptoms, emotion regulation, and psychosocial functioning in community veterans. Peer-supported, transdiagnostic mHealth programs may be a particularly efficient, effective, and low-burden approach to improving mental health among trauma-exposed populations. Investigation of peer-supported programs among other populations is necessary to evaluate the generalizability of the findings. Analyses comparing peer support that was required versus optional indicated that some veterans may not need or want peer support. Future research should evaluate how best to deliver peer support and for whom it is most beneficial. If successful, peer-supported tech programs may increase the Veteran Affairs workforce as well as improve veteran mental health services and outcomes. ClinicalTrials.gov NCT04286165; https://clinicaltrials.gov/study/NCT04286165.

Investigating the Potential of Multilineage Differentiating Stress-Enduring Cells for Osteochondral Healing.

Multilineage differentiating stress-enduring (Muse) cells, a pluripotent stem cell subset of mesenchymal stem cells (MSCs), have shown promise for various tissue repairs due to their stress tolerance and multipotent capabilities. We aimed to investigate the differentiation potential in vitro, the dynamics in vivo, and the reparative contribution of Muse cells to osteochondral lesions. Labeled MSCs were cultured and sorted into Muse and non-Muse (MSCs without Muse cells) groups. These cells were then formed into spheroids, and chondrogenic differentiation was assessed in vitro. Twenty-one immunocompromised mice were used as the in vivo models of osteochondral lesions. Live imaging, macroscopic evaluation, and histological and immunohistochemical analyses were conducted at the 4- and 8-week time points. Muse cell spheroids were formed, which were larger and stained more intensely with toluidine blue than non-Muse spheroids, indicating better chondrogenic differentiation. Live imaging confirmed luminescence in all 4-week model knees, but only in a few knees at 8 weeks, suggesting cell persistence. Macroscopically and histologically, no significant differences were observed between the Muse and non-Muse groups at 4 and 8 weeks; however, both groups showed better cartilage repair than that of the vehicle group at 8 weeks. No collagen type II generation was observed in the repaired tissues. The implantation of the spheroids of Muse and non-Muse cells resulted in better healing of osteochondral lesions than that of the controls, and Muse cells had a higher chondrogenic differentiation potential in vitro than non-Muse cells.

Longitudinal in vivo cationic contrast-enhanced computed tomography classifies equine articular cartilage injury and repair.

Cationic contrast-enhanced computed tomography (CECT) capitalizes on increased contrast agent affinity to the charged proteoglycans in articular cartilage matrix to provide quantitative assessment of proteoglycan content with enhanced images. While high resolution microCT has demonstrated success, we investigate cationic CECT use in longitudinal in vivo imaging at clinical resolution. We hypothesize that repeated administration of CA4+ will have no adverse side effects or complications, and that sequential in vivo imaging assessments will distinguish articular cartilage repair tissue from early degenerative and healthy cartilage in critically sized chondral defects. In an established equine translational preclinical model, lameness and synovial effusion scores are similar to controls after repeated injections of CA4+ (eight injections over 16 weeks) compared to controls. Synovial fluid total protein, leukocyte concentration, and sGAG and PGE2 concentrations and articular cartilage and synovial membrane scores are also equivalent to controls. Longitudinal in vivo cationic CECT attenuation in repair tissue is significantly lower than peripheral to (adjacent) and distantly from defects (remote sites) by 4 weeks (p < 0.001), and this difference persists until 16 weeks. At the 6- and 8-week time points, the adjacent locations exhibit significantly lower cationic CECT attenuation compared with the remote sites, reflecting peri-defect degeneration (p < 0.01). Cationic CECT attenuation at clinical resolution significantly correlates with cationic CECT (microCT) (r = 0.69, p < 0.0001), sGAG (r = 0.48, p < 0.0001), and ICRS II histology score (r = 0.63, p < 0.0001). In vivo cationic CECT imaging at clinical resolution distinguishes fibrous repair tissue from degenerative and healthy hyaline cartilage and correlates with molecular tissue properties of articular cartilage.

Longitudinal analysis of host protein serum signatures of treatment and recovery in pulmonary tuberculosis.

A better understanding of treatment progression and recovery in pulmonary tuberculosis (TB) infectious disease is crucial. This study analyzed longitudinal serum samples from pulmonary TB patients undergoing interventional treatment to identify surrogate markers for TB-related outcomes. Serum that was collected at baseline and 8, 17, 26, and 52 weeks from 30 TB patients experiencing durable cure were evaluated and compared using a sensitive LC-MS/MS proteomic platform for the detection and quantification of differential host protein signatures relative to timepoint. The global proteome signature was analyzed for statistical differences across the time course and between disease severity and treatment groups. A total of 676 proteins showed differential expression in the serum over these timepoints relative to baseline. Comparisons to understand serum protein dynamics at 8 weeks, treatment endpoints at 17 and 26 weeks, and post-treatment at 52 weeks were performed. The largest protein abundance changes were observed at 8 weeks as the initial effects of antibiotic treatment strongly impacted inflammatory and immune modulated responses. However, the largest number of proteome changes was observed at the end of treatment time points 17 and 26 weeks respectively. Post-treatment 52-week results showed an abatement of differential proteome signatures from end of treatment, though interestingly those proteins uniquely significant at post-treatment were almost exclusively downregulated. Patients were additionally stratified based upon disease severity and compared across all timepoints, identifying 461 discriminating proteome signatures. These proteome signatures collapsed into discrete expression profiles with distinct pathways across immune activation and signaling, hemostasis, and metabolism annotations. Insulin-like growth factor (IGF) and Integrin signaling maintained a severity signature through 52 weeks, implying an intrinsic disease severity signature well into the post-treatment timeframe. Previous proteome studies have primarily focused on the 8-week timepoint in relation to culture conversion status. While this study confirms previous observations, it also highlights some differences. The inclusion of additional end of treatment and post-treatment time points offers a more comprehensive assessment of treatment progression within the serum proteome. Examining the expression dynamics at these later time periods will help in the investigation of relapse patients and has provided indicative markers of response and recovery.

Scaffold-Free Bone Marrow-Derived Mesenchymal Stem Cell Sheets Enhance Bone Formation in a Weight-Bearing Rat Critical Bone Defect Model.

Researchers have been exploring alternative methods for bone tissue engineering, as current management of critical bone defects may be a significant challenge for both patient and surgeon with conventional surgical treatments associated with several potential complications and drawbacks. Recent studies have shown mesenchymal stem cell sheets may enhance bone regeneration in different animal models. We investigated the efficacy of implanted scaffold-free bone marrow-derived mesenchymal stem cell (BMSC) sheets on bone regeneration of a critical bone defect in a weight-bearing rat model. BMSCs were isolated from the femora of male Sprague-Dawley rats 5-6 weeks of age and cell sheets were produced on temperature-responsive culture dishes. Nine male Sprague-Dawley rats 6-8 weeks of age were utilized. A bilateral femoral critical bone defect was created with a bridge plate serving as internal fixation. One side was randomly selected and BMSC sheets were implanted into the bone defect (BMSC group), with the contralateral side receiving no treatment (control). Rats were anesthetized and radiographs were performed at 2-week intervals. At the 8-week time point, rats were euthanized, femurs harvested, and microcomputed tomography and histological analysis was performed. We found a statistically significant increase in new bone formation and bone volume fraction compared with the control. Histomorphometry analysis revealed a larger percent of newly formed bone and a higher total histological score. Our results suggest that scaffold-free BMSC sheets may be used in the management of large weight-bearing bone defects to complement a different surgical technique or as a standalone approach followed by internal fixation. However, further research is still needed.

Home-Based Physical Activity Program With Health Coaching for Participants With Chronic Obstructive Pulmonary Disease in Sweden: A Proof-of-Concept Pilot Study

Home-based interventions are at the center stage of current health care demands. There is a clear need to translate pulmonary rehabilitation into a home-based setting. This 8-week pilot study aimed to determine the feasibility of a home-based physical activity program for participants with chronic obstructive pulmonary disease (COPD) in Sweden. Patients with COPD, aged 40 years or older and clinically stable in the past 3 months, were recruited. The program used a fitness tracker to monitor step count, weekly health coaching calls using motivational interviewing, and video-guided mindful movements. The outcome measures were adherence to the 8-week program’s video-guided exercises (number of times videos were watched), adherence to health coaching calls (minimum 8), monthly and daily step count, and quality of life (QoL) using the chronic respiratory questionnaire. Thirteen participants were enrolled, and 12 participants adhered to health coaching calls and step monitoring. We had 643 video-exercise views, which exceeded the minimum standard (576 views). The mean difference comparing total monthly steps from baseline and the 8-week time point was 47,039 steps (95% CI, –113,625 to 1623.5; P=.06). The minimal clinical improvement of 500 daily steps was found for 8 of the patients. No significant improvement was found in the QoL measures and mental health. We found the home-based physical activity program to be a feasible intervention. Patients reported high adherence to tracking step counts, health coaching calls, and video-guided exercise. No improvements in QoL or monthly step count emerged; however, we found high adherence and a positive trend in the number of monthly step counts, and improvements of at least 500 daily step counts improved in most patients with this small sample size.

Perineurial Window is Critical for Experimental Reverse End-to-Side Nerve Transfer.

The depth of connective tissue window in the side of a recipient nerve in reverse end-to-side transfers (RETS) remains controversial. To test whether the depth of connective tissue disruption influences the efficiency of donor axonal regeneration in the context of RETS. Sprague-Dawley rats (n = 24) were assigned to 1 of the 3 groups for obturator nerve to motor femoral nerve RETS: group 1, without epineurium opening; group 2, with epineurium only opening; and group 3, with epineurium and perineurium opening. Triple retrograde labeling was used to assess the number of motor neurons that had regenerated into the recipient motor femoral branch. Thy1-GFP rats (n = 8) were also used to visualize the regeneration pathways in the nerve transfer networks at 2- and 8-week time point using light sheet fluorescence microscopy. The number of retrogradely labeled motor neurons that had regenerated distally toward the target muscle was significantly higher in group 3 than that in groups 1 and 2. Immunohistochemistry validated the degree of connective tissue disruption among the 3 groups, and optical tissue clearing methods demonstrated donor axons traveling outside the fascicles in groups 1 and 2 but mostly within the fascicles in group 3. Creating a perineurial window in the side of recipient nerves provides the best chances of robust donor axonal regeneration across the RETS repair site. This finding aids nerve surgeons by confirming that a deep window should be undertaken when doing a RETS procedure.

Successful retrograde regeneration using a sensory branch for motor nerve transfer.

The objective of this study was to test whether regenerating motor axons from a donor nerve can travel in a retrograde fashion using sensory branches to successfully reinnervate a motor nerve end organ. This study has two parts. In part I, rats (n = 30) were assigned to one of five groups for obturator nerve (ON)-to-femoral nerve transfer: group 1, ON-to-saphenous nerve (SN) distal stump; group 2, ON-to-SN proximal stump without femoral nerve proper (FNP) injury; group 3, ON-to-SN proximal stump with FNP crush injury; group 4, ON-to-SN proximal stump with FNP transection injury; and group 5, gold standard transfer, ON-to-motor femoral nerve (MFN) branch. At 8 weeks, retrograde labeling was done from the distal MFN, and the spinal cords were examined to assess the degree of obturator motor axon regeneration across the five groups. In part II, only group 4 was examined (n = 8). Through use of immunostaining and optical tissue clearing methods, the nerve transfer networks were cleared and imaged using light-sheet fluorescence microscopy to visualize the regeneration pathways in 2D and 3D models at 2- and 8-week time points. Proximal FNP transection (group 4) enabled a significantly higher number of retrogradely regenerated motor axons compared with control groups 1-3. Moreover, group 4 had modest, but nonsignificant, superiority of motor neuron counts compared with the positive control group, group 5. Optical tissue clearing demonstrated that the axons traveled in a retrograde fashion from the recipient sensory branch to the FNP mixed stump, then through complex turns, down to distal branches. Immunostaining confirmed the tissue clearing findings and suggested perineurium disruption as a means by which axons could traverse across fascicular boundaries. Sensory branches can transmit regenerating axons from donor nerves back to main mixed recipient nerves, then distally toward target organs. The extent of retrograde regeneration is markedly influenced by the type and severity of injury sustained by the recipient nerve. Using a sensory branch as a bridge for retrogradely regenerating axons can open new potential horizons in nerve repair surgery for severely injured mixed nerves.

Inhibition of Hypoxia Inducible Factor Prolyl Hydroxylase By FG-4592 (Roxadustat) Increases Erythropoietic Stimulation in a Mouse Model of Sickle Cell Disease

Sickle cell disease (SCD) is a devastating monogenic disorder caused by a mutation in the β-globin gene, leading to production of mutant hemoglobin S (HbS). Abnormal polymerization of deoxygenated HbS causes sickling of red blood cells (RBCs), resulting in painful vaso-occlusive crises and chronic hemolytic anemia. Etiologies of chronic anemia in SCD are multifactorial, including intravascular and extravascular hemolysis, inadequate erythropoietin (EPO) production, and abnormal iron homeostasis (Xu and Thein, Blood 2022). Hydroxyurea (HU) is the standard-of-care drug therapy for SCD and primarily reduces HbS polymerization through induction of fetal Hb (HbF) expression. However, it only modestly improves anemia and does not address other causes of anemia in SCD. Additionally, lower Hb levels in SCD are associated with chronic kidney disease (CKD), stroke, and pulmonary hypertension, arguing for the need for additional therapies targeting anemia in SCD. Hypoxia-inducible factor prolyl hydroxylase inhibitors (HIF-PHIs) are a class of drugs developed for treatment of anemia in CKD and end-stage renal disease. HIF-PHIs stimulate erythropoiesis by inhibiting prolyl hydroxylase and von Hippel-Lindau (VHL)-mediated degradation of HIF-α, leading to increased hypoxic signaling and promotion of HIF-induced EPO expression in the kidneys and liver. Stimulation of erythropoiesis also increases production of erythroferrone, the main regulator of hepcidin, and suppression of hepcidin levels by HIF-PHIs may improve iron mobilization. Furthermore, HIF-PHIs can induce HbF expression in erythroblasts derived from CD34+ hematopoietic stem and progenitor cells (HSPCs) (Hsieh et al.,Blood 2007, Feng et al., ASH abstract 2021), suggesting multiple potential therapeutic benefits in SCD. Therefore, we characterized the effect of the HIF-PHI FG-4592 (roxadustat), on HbF induction in CD34+ HSPCs and on hematologic markers, EPO and hepcidin expression in a mouse model of SCD. CD34+ HSPCs from 3 donors were differentiated in vitro to erythroblasts with vehicle, 20 μM of FG-4592, or 50 μM of HU. Flow cytometric analysis on Day 9 showed that FG-4592 treatment increased % F-cells by 1.47-fold (±0.45), compared to a 2.21-fold (±0.89) increase with HU (Fig. 1). We then transplanted lethally irradiated (1000 rads) male 6-8-week-old C57BL/6J mice with bone marrow from transgenic sickle cell mice (Hb SS; Townes model, Jackson Laboratory). Twelve weeks after transplantation, mice with >80% engraftment of the donor HbSS phenotype, determined by Hb electrophoresis, were used for subsequent experiments. Vehicle, 25 mg/kg FG-4592, or 50 mg/kg FG-4592 was administered three times weekly through intraperitoneal injection for 8 weeks. All mice underwent retro-orbital bleeding at baseline (48 h pretreatment), 2, 4, 6, and 8 weeks of treatment for hematological evaluation, including Hb, hematocrit, and reticulocyte % (Fig. 1). At the end of 8 weeks, livers, kidneys, and plasma were collected. Hb and hematocrit levels increased in the 50 mg/kg FG-4592 group compared to the other groups at 4 weeks but decreased by 8 weeks for all groups. Reticulocyte % also decreased in all groups at the 4- and 8-week timepoints. Red cell distribution width (RDW) and platelet count up-trended while mean corpuscular volume (MCV) and mean corpuscular Hb (MCH) down-trended over the 8 weeks, suggestive of an evolving iron deficiency contributing to worsening anemia. Quantitative PCR detected a 182% increase in liver EPO transcript and a 68% increase in kidney EPO transcript in the 50 mg/kg FG-4592 treated group, compared to vehicle. No changes in liver hepcidin transcription were observed in any group. ELISA analysis detected increased plasma EPO and decreased plasma hepcidin levels with increasing doses of FG-4592. None of these trends, except increases in RDW, were statistically significant. Our study is the first to characterize the effect of HIF-PHI drugs in a mouse model of SCD. Preliminary data confirm an effect of FG-4592 on HbF induction and suggest that FG-4592 may increase EPO expression and decrease hepcidin levels in SCD mice. These results highlight the potential of prolyl hydroxylase inhibition as a novel therapeutic approach for chronic anemia in SCD. Future directions include testing higher doses of FG-4592 to induce a stronger erythropoietic effect and monitoring additional markers of iron homeostasis in our mouse model of SCD. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

P9. Sex differences in response to recombinant human bone morphogenetic protein-2 (rhBMP-2) in a rat posterolateral fusion spine model

<h3>BACKGROUND CONTEXT</h3> Estrogen modulates bone activity largely through inhibition of osteoclast bone resorption and bone morphogenetic protein-2 (BMP-2) signaling pathways in osteoblasts. Minimal data exist concerning potential sex-dependent differences in BMP-2-mediated bone regeneration. Here, we aimed to quantify sex-dependent differences in the bone healing response to recombinant human BMP-2 (rhBMP-2) treatment in a rat posterolateral spinal fusion model. <h3>METHODS</h3> A total of 48 female and male Sprague-Dawley rats (N=24/group), underwent L4-L5 posterolateral fusion with bilateral placement of absorbable collagen sponges, loaded with 5 µg of rhBMP-2 (10 µg/animal). At 8 weeks postoperatively, 10 specimens of each sex were tested in flexion-extension with quantification of range of motion (ROM) and stiffness. The remaining specimens were evaluated for new bone growth and fusion via radiography, blinded manual palpation and microcomputed tomography (microCT). Laboratory microCT quantified bone microarchitecture and synchrotron microCT examined bone microstructure. On histology, the number of adipocytes within a standardized field of view was counted for both male and female fusion masses. <h3>RESULTS</h3> Manual palpation scores differed significantly between sexes, with mean fusion scores of 2.4±0.4 in females vs 3.1±0.6 in males, p < 0.001. Biomechanical stiffness did not differ between sexes, but ROM was significantly greater and more variable for females vs males (3.7°±5.6° vs 0.27°±0.15°,p < 0.005, respectively. Laboratory microCT showed significantly smaller volumes of fusion masses in females vs males (262±87 mm³ vs 732±238 mm³, respectively, p < 0.001) but significantly higher bone volume fraction (0.27±0.08 vs 0.12±0.05, respectively, p < 0.001). Mean trabecular thickness was not different, but trabecular number was significantly greater in females (3.1±0.5 mm-1vs 1.5±0.4 mm-1, respectively, p<0.001). Synchrotron microCT showed fine bone structures developing in both sexes at the 8-week time point. Adipocyte density was found to be significantly higher in male fusion masses than in female fusion masses. <h3>CONCLUSIONS</h3> This study demonstrates that sex-based factors may influence both the quantity and quality of bone formation in patients receiving rhBMP-2 for a variety of orthopedic applications. We found that male rats had significantly higher fusion scores, greater fusion-mass volume, and lower ratio of new bone volume to total volume compared to female rats. Male and female mean trabecular thicknesses were equal, but the female fusion masses were more densely filled with trabeculae (higher trabecular number) than those of males. MicroCT results suggested that the male trabeculae were more rod-like and the female more plate-like, although the methods that were used are generally considered less accurate for trabecular bone, which has a high degree of concavity. The incomplete fusions on three female rats, evidenced by high ROM and small gaps on microCT, suggest that females may have a more variable response to rhBMP-2. Investigation into whether these sex-based differences are specific to rhBMP-2-induced bone formation or are more general to bone regeneration/healing is prudent. <h3>FDA DEVICE/DRUG STATUS</h3> This abstract does not discuss or include any applicable devices or drugs.

Effect of Compression Therapy in the Treatment of Tibial Stress Syndrome in Military Service Members.

Tibial stress syndrome (TSS) is an overuse injury of the lower extremities. There is a high incidence rate of TSS among military recruits. Compression therapy is used to treat a wide array of musculoskeletal injuries. The purpose of this study was to investigate the use of compression therapy as a treatment for TSS in military service members. A parallel randomized study design was utilized. Military members diagnosed with TSS were assigned to either a relative rest group or compression garment group. Both groups started the study with 2 weeks of lower extremity rest followed by a graduated running program during the next 6 weeks. The compression garment group additionally wore a shin splints compression wrap during the waking hours of the first 2 weeks and during activity only for the next 6 weeks. Feelings of pain, TSS symptoms, and the ability to run 2 miles pain free were assessed at baseline, 4 weeks, and 8 weeks into the study. Feelings of pain and TSS symptoms decreased during the 8-week study in both groups (P < .05), but these changes were not significantly different between groups (P > .05). The proportion of participants who were able to run 2 miles pain free was significantly different (P < .05) between the 2 groups at the 8-week time point with the compression garment group having a significantly increased ability to complete the run without pain. Although perceptions of pain at rest were not different between groups, the functional ability of running 2 miles pain free was significantly improved in the compression garment group. These findings suggest that there is a moderate benefit to using compression therapy as an adjunct treatment for TSS, promoting a return to training for military service members.

Cardioprotective Effects of Preconditioning Exercise in the Female Tumor‐Bearing Mouse

BackgroundCancer cachexia, a metabolic wasting syndrome, affects up to 80% of cancer patients and leads to the death in up to 20% of cancer patients. While research is growing in the field, there are still no clear diagnostic criteria and cancer cachexia remains an untreated condition. Aerobic exercise has been shown to positively impact cachexia by slowing its development and attenuating muscle loss. The most effective timing, duration, and intensity of exercise as a preventative and protective measure against cancer cachexia remains questionable. Therefore, the purpose of this study was to examine the effects of preconditioning exercise as a protective measure for tumor‐mediated muscle wasting.MethodsFemale LC3 Tg+ and WT mice were randomly separated into four groups, sedentary non‐tumor bearing (SED+NT), sedentary tumor bearing (SED+T), treadmill exercise non‐tumor bearing (TM+NT), and treadmill exercise tumor bearing (TM+T). Mice underwent an 8‐week treadmill exercise training protocol (TM) or remained sedentary (SED). Next, mice were implanted with tumor cells (T group; 1x105 LLC cells in flank) or remained non‐tumor (NT) for 4 weeks. To examine the protective effects of exercise on tumor growth and muscle wasting during tumor‐mediated muscle wasting, grip strength and tumor growth evaluations were taken at baseline, 8‐week, and 12‐week time points. At the end of the 12‐week study, muscle and tumor tissue was collected and weighed.ResultsTumor bearing resulted in a significant decline in cardiac function. SED+T showed a significant decrease in fractional shortening (P &lt; 0.05) when compared to the other groups. This coincided with an increase in beclin‐1 expression in SED+T mice. Interestingly, preconditioning exercise (exercise prior to tumor bearing) appeared to preserve cardiac function (TM+T not significantly different than SED+NT). Exercise‐mediated cardioprotection also coincided with abolished beclin‐1 signaling since this was not significantly elevated in TM+T mice. Additionally, TM resulted in a 20‐fold decrease in estimated tumor volume (P &lt; 0.05) and a 60% decrease in tumor mass (P &lt; 0.05) compared to SED tumors.ConclusionThe data indicate potential cardioprotective effects of preconditioning exercise on preserving cardiac structure and function, as well as regulating autophagic pathways (beclin‐1) during tumor bearing. Preconditioning exercise may be an effective and accessible treatment intervention for early‐stage cancer survivors. This data is crucial in identifying the significance of exercise and the timing of exercise as a protective measure against the detrimental effects of cancer cachexia.

Effectiveness and Safety of Fecal Microbiota Transplantation for Clostridioides Difficile Infection: Results From a 5344-Patient Cohort Study

Effectiveness and Safety of Fecal Microbiota Transplantation for Clostridioides Difficile Infection: Results From a 5344-Patient Cohort Study

1039. Rapid Restoration of Bile Acid Compositions After Treatment with RBX2660 for Recurrent Clostridioides difficile Infection—Results from the PUNCH CD3 Phase 3 Trial

BackgroundMicrobiota-based treatments are increasingly evaluated as a strategy to reduce recurrence of Clostridioides difficile infection (rCDI), and their proposed mechanisms include restoration of the microbiota and microbiota-mediated functions, including bile acid metabolism. RBX2660—a broad-consortium investigational live biotherapeutic—has been evaluated in >600 participants in 6 clinical trials, with consistent reduction of rCDI recurrence. Here we report that fecal bile acid compositions were significantly restored in treatment-responsive participants in PUNCH CD3—a Phase 3 randomized, double-blinded, placebo-controlled trial of RBX2660.MethodsPUNCH CD3 participants received a single dose of RBX2660 or placebo between 24 to 72 hours after completing rCDI antibiotic treatment. Clinical response was the absence of CDI recurrence at eight weeks after treatment. Participants voluntarily submitted stool samples prior to blinded study treatment (baseline), 1, 4 and 8 weeks, 3 and 6 months after receiving study treatment. A liquid chromatography tandem mass spectrometry method was developed to extract and quantify 33 bile acids from all participant fecal samples received up to the 8-week time point. Mean bile acid compositions were fit to a Dirichlet multinomial distribution and compared across time points and between RBX2660- and placebo-treated participants.ResultsClinically, RBX2660 demonstrated superior efficacy versus placebo (70.4% versus 58.1%). RBX2660-treated clinical responders’ bile acid compositions shifted significantly from before to after treatment. Specifically, primary bile acids predominated before treatment, whereas secondary bile acids predominated after treatment (Figure 1A). These changes trended higher among RBX2660 responders compared to placebo responders. Importantly, median levels of lithocholic acid (LCA) and deoxycholic acid (DCA) showed large, significant increases after treatment (Figure 1B). A. Bile acid compositions before (BL) and up to 8 weeks after RBX2660 treatment among treatment responders. Compositions are shown as the fraction of total bile acids classified as primary or secondary conjugated or deconjugated bile acids. B. Concentrations of lithocholic acid (LCA) and deoxycholic acid (DCA) among RBX2660 treatment responders, shown with individual samples and time point group median with interquartile ranges.ConclusionAmong PUNCH CD3 clinical responders, RBX2660 significantly restored bile acids from less to more healthy compositions. These clinically correlated bile acid shifts are highly consistent with results from a prior trial of RBX2660.Disclosures Romeo Papazyan, PhD, Ferring Research Institute (Employee) Bryan Fuchs, PhD, Ferring Pharmaceuticals (Employee) Ken Blount, PhD, Rebiotix Inc., a Ferring Company (Employee)

The impact of internet-based cognitive behaviour therapy for perfectionism on different measures of perfectionism: a randomised controlled trial

ABSTRACT The current study investigated the impact of an 8-module internet-based cognitive behaviour therapy for perfectionism (ICBT-P) across a variety of perfectionism subscales. Undergraduate students who identified as having a problem with perfectionism were randomized to receive the intervention (n = 41), and were free to choose the number of treatment modules they completed over a 4-week period, while the control group (N = 48) received access to treatment 8 weeks post-randomisation. Secondary measures included depression, anxiety, stress, body image and self-compassion. Assessments occurred at baseline, 2-, 4- and 8-week time points. A mean of 3.12 (SD = 2.67) modules were completed; 7 participants (17%) completed none and 6 (15%) completed all. Linear mixed modelling (with baseline observation included as a covariate) showed significant Bonferroni-adjusted post-hoc between-group differences for 5 of the 6 perfectionism measures, favouring the intervention group; the most robust between group effect sizes were for the Concern over Mistakes (d = −0.82), High Standards (d = −0.69), and Perfectionistic Standards (d = −0.47) subscales. There were no between-group differences for our secondary measures. ICBT-P was found to be an effective intervention for reducing different components of perfectionism compared to a control group. The relatively low use of modules may have contributed to a lack of effect on secondary measures. Australian New Zealand Clinical Trials Registry (ANZCTR) Trial Number: ACTRN12620000562976

Protective Effects of Preconditioning Exercise on Skeletal Muscle Strength and Tumor Growth in the Female Tumor‐Bearing Mouse

Background Cancer cachexia, a complex metabolic wasting disease, occurs in up to 80% of cancer patients and results in death in up to one-third of cancer patients. While research in this field is growing, cancer cachexia remains an untreated condition with irreversible effects as there are no clear diagnostic criteria. Therefore, a great need exists to better understand this disease and possible preventative methods. Research has shown that exercise interventions have a positive impact on inhibiting tumor growth and preserving muscle mass. Many questions remain regarding the most effective timing and extent of exercise as a protective measure. Therefore, the purpose of this study was to examine the effects of preconditioning exercise as a protective measure for cancer cachexia. Methods Female LC3 Tg+ and WT mice were randomly separated into four groups, sedentary non-tumor bearing (SED+NT), sedentary tumor bearing (SED+T), treadmill exercise non-tumor bearing (TM+NT), and treadmill exercise tumor bearing (TM+T). Mice underwent an 8-week treadmill exercise training protocol (TM) or remained sedentary (SED). Next, mice were implanted with tumor cells (T group; 1x105 LLC cells in flank) or remained non-tumor (NT) for 4 weeks. To examine the protective effects of exercise on tumor growth and muscle wasting during cancer cachexia, grip strength and tumor growth evaluations were taken at baseline, 8-week, and 12-week time points. At the end of the 12-week study, muscle and tumor tissue was collected and weighed. Results While no differences existed between groups at baseline, exercise trained (TM) mice exhibited modest gains in grip strength compared to SED mice. Following tumor bearing, SED+T mice exhibited a significant (P < 0.05) decline in grip strength compared to SED+NT animals. Additionally, SED+T skeletal muscle mass (gastrocnemius, mixed fiber) was significantly (P < 0.05) less compared to TM+T. Interestingly, TM resulted in a 20-fold decrease in estimated tumor volume (P < 0.05) and a 60% decrease in tumor mass (P < 0.05) compared to SED tumors. Conclusion The analyzed data indicate that preconditioning exercise appears to play an important role in protecting the musculature during cancer cachexia. This is evidenced by exercise's ability to guard against cancer cachexia-mediated declines in both muscle strength and mass. Furthermore, exercise preconditioning significantly stunted tumor growth even when animals did not exercise during the tumor bearing. This information is crucial in understanding the significance of exercise in cancer patients and in elucidating the importance of timing of exercise as a protective measure against the detrimental effects of cancer cachexia.

Cerebrovascular Neuroprotection after Acute Concussion in Adolescents.

To assess acute cerebrovascular function in concussed adolescents (14-21 years of age), whether it is related to resting cerebral hemodynamics, and whether it recovers chronically. Cerebral vasoreactivity and autoregulation, based on middle cerebral artery blood flow velocity, was assessed in 28 concussed participants (≤14 days of injury) and 29 matched controls. The participants in the concussion group returned for an 8-week follow-up assessment. Over the course of those 8-weeks, participants recorded aerobic exercise frequency and duration. Between groups, demographic, clinical, and hemodynamic variables were not significantly different. Vasoreactivity was significantly higher in the concussed group (p=0.02). Within the concussed group, 60% of the variability in resting cerebral blood flow velocity was explained by vasoreactivity and two components of autoregulation - falling slope and effectiveness of autoregulation (adjusted R2 =0.60, p < 0.001). Moreover, lower mean arterial pressure, lower responses to increases in arterial pressure, and lower vasoreactivity were significantly associated with larger symptom burden (adjusted R2 =0.72, p < 0.01). By the 8-week timepoint, symptom burden, but not vasoreactivity, improved in all but four concussed participants (p < 0.01). 8-week change in vasoreactivity was positively associated with aerobic exercise volume (adjusted R2 =0.19, p=0.02). Concussion resulted in changes in cerebrovascular regulatory mechanisms, which in turn explained the variability in resting cerebral blood flow velocity and acute symptom burden. Furthermore, these alterations persisted chronically despite symptom resolution, but was positively modified by aerobic exercise volume. These findings provide a mechanistic framework for further investigation into underlying cerebrovascular related symptomatology. ANN NEUROL 2021;90:43-51.

Articular cartilage regeneration in rats using human bone marrow stromal cells

Purpose: Injury to articular cartilage found in knee joints is often disabling and affects both younger and older individuals. Injured cartilage rarely heals, leading to a progressive loss of normal function of the joint in many cases. Current treatments are far from satisfactory. One treatment is surgery (such as drilling through the cartilage to the bone, abrasion to remove diseased cartilage and microfracture). These procedures can result in the formation of a mixture of fibrous tissue and cartilage within the injury, which does not lead to a total recovery of joint function. Autologous human bone marrow stromal cell (hBMSC) transplantation maybe a potential avenue to regenerate articular cartilage. We previously determined that naive (untreated) hBMSC were able to form stable, non-hypertrophic cartilage when transplanted subcutaneously in conjunction with fibrin microbeads covalently coated with hyaluronic acid (FMBs). In our current study, we are investigating the ability of hBMSC/FMB constructs to generate native-like cartilage in an articular defect in immunocompromised rodents. Methods: hBMSCs were obtained from surgical bone waste and grown in culture for 2-3 passages. Approval for this study was given by the institutional Animal Care and Use Committee (ACUC). To create the articular cartilage defects in immunocompromised SRG rats (Hera/Charles River), the patella was laterally displaced and the distal femur was immobilized using a clamp. The defect was made in the trochlear groove of the distal femur using a 1.8mm diameter microdrill. The defect was either left unrepaired (sham), or subsequently filled with either FMBs alone (FMBs Alone), hBMSCs attached to FMBs (FMBs+cells), or with a pellet created by incubating BMSCs with FMBs for 10 days in chondrogenic medium. At the 4-week and 8-week timepoints following surgery, the rats were euthanized and femurs were harvested and fixed in 4% paraformaldehyde for 24 hours and then decalcified using a 10% EDTA solution for 30 days. After decalcification, the femurs were paraffin embedded and sectioned at a thickness of 6 μm. Histological staining was performed using hematoxylin and eosin, toluidine blue, and safranin O methods to assess cartilage formation. Immunohistochemistry for aggrecan within the cartilage was also performed to assess cartilage proteoglycan formation. Image analysis using Image J was performed on sections stained with safranin O. Cartilage thickness was approximated based on the thickness of the intact cartilage surrounding the defect, and a region of interest (ROI) was drawn surrounding the defect with the same depth as the intact cartilage. The image color channels were separated into red, green, and blue, and the blue channel was used to threshold for the stained proteoglycan. The “analyze particles” module of Image J was used to calculate the percentage of the ROI that stained positive for proteoglycan. Second-harmonic generation microscopy was also used to evaluate collagen fibril structure and organization. Results: Toluidine blue staining revealed cartilage formation in defects at the 4-week and 8-week timepoints in both the FMBs+cells and pellet groups. No cartilage was detected in the FMBs alone or sham groups (Figure 1). Percentages of defect area positive for cartilage proteoglycan in the FMBs+cells, pellet, FMBs alone, and sham group defect areas were 8.95%, 3.03%, 0.008%, and 0.094% respectively. A comparable ROI in healthy rat cartilage showed proteoglycan positivity of 58.43% of the area (Figure 2). Second harmonic generation imaging of the defect area showed that from sham to FMB alone to pellet to FMBs+cells groups, the organization of the collagen fibrils begin to more closely approximate healthy rat cartilage (Figure 3). Long-term time points are currently being analyzed. Conclusions: At the early timepoints presented in this abstract, there is evidence of cartilage formation in the drilled articular cartilage defects in both the FMBs+cells and pellet groups. This initial data shows the potential for our method to regenerate articular cartilage in vivo. Analysis of our long-term time points will reveal the extent and quality of healing induced by hBMSCs transplanted in conjunction with FMBs.

Generation of Self-assembled Vascularized Human Skin Equivalents.

Human skin equivalents (HSEs) are tissue engineered constructs that model epidermal and dermal components of human skin. These models have been used to study skin development, wound healing, and grafting techniques. Many HSEs continue to lack vasculature and are additionally analyzed through post-culture histological sectioning which limits volumetric assessment of the structure. Presented here is a straightforward protocol utilizing accessible materials to generate vascularized human skin equivalents (VHSE); further described are volumetric imaging and quantification techniques of these constructs. Briefly, VHSEs are constructed in 12 well culture inserts in which dermal and epidermal cells are seeded into rat tail collagen type I gel. The dermal compartment is made up of fibroblast and endothelial cells dispersed throughout collagen gel. The epidermal compartment is made up of keratinocytes (skin epithelial cells) that differentiate at the air-liquid interface. Importantly, these methods are customizable based on needs of the researcher, with results demonstrating VHSE generation with two different fibroblast cell types: human dermal fibroblasts (hDF) and human lung fibroblasts (IMR90s). VHSEs were developed, imaged through confocal microscopy, and volumetrically analyzed using computational software at 4- and 8-week timepoints. An optimized process to fix, stain, image, and clear VHSEs for volumetric examination is described. This comprehensive model, imaging, and analysis techniques are readily customizable to the specific research needs of individual labs with or without prior HSE experience.

A Novel Strategy to Enhance Microfracture Treatment With Stromal Cell-Derived Factor-1 in a Rat Model.

BackgroundMicrofracture is one of the most widely used techniques for the repair of articular cartilage. However, microfracture often results in filling of the chondral defect with fibrocartilage, which exhibits poor durability and sub-optimal mechanical properties. Stromal cell-derived factor-1 (SDF-1) is a potent chemoattractant for mesenchymal stem cells (MSCs) and is expressed at high levels in bone marrow adjacent to developing cartilage during endochondral bone formation. Integrating SDF-1 into an implantable collagen scaffold may provide a chondro-conductive and chondro-inductive milieu via chemotaxis of MSCs and promotion of chondrogenic differentiation, facilitating more robust hyaline cartilage formation following microfracture.ObjectiveThis work aimed to confirm the chemoattractive properties of SDF-1 in vitro and develop a one-step method for incorporating SDF-1 in vivo to enhance cartilage repair using a rat osteochondral defect model.MethodsBone marrow-derived MSCs (BMSCs) were harvested from the femurs of Sprague–Dawley rats and cultured in low-glucose Dulbecco’s modified Eagle’s medium containing 10% fetal bovine serum, with the medium changed every 3 days. Passage 1 MSCs were analyzed by flow cytometry with an S3 Cell Sorter (Bio-Rad). In vitro cell migration assays were performed on MSCs by labeling cells with carboxyfluorescein diacetate, succinimidyl ester (CFDA-SE; Bio-Rad). For the microfracture model, a 1.6-mm-diameter osteochondral defect was created in the femoral trochleae of 20 Sprague–Dawley rats bilaterally until bone marrow spillage was seen under saline irrigation. One knee was chosen at random to receive implantation of the scaffold, and the contralateral knee was left unfilled as an empty control. Type I collagen scaffolds (Kensey Nash) were coated with either gelatin only or gelatin and SDF-1 using a dip coating process. The rats received implantation of either a gelatin-only scaffold (N = 10) or gelatin-and-SDF-1 scaffold (N = 10) at the site of the microfracture. Femurs were collected for histological analyses at 4- and 8-week time points post-operatively, and sections were stained with Safranin O/Fast Green. The samples were graded blindly by two observers using the Modified O’Driscoll score, a validated scoring system for chondral repair. A minimum of 10 separate grading scores were made per sample and averaged. Quantitative comparisons of cell migration in vitro were performed with one-way ANOVA. Cartilage repair in vivo was also compared among groups with one-way ANOVA, and the results were presented as mean ± standard deviation, with P-values < 0.05 considered as statistically significant.ResultsMSC migration showed a dose–response relationship with SDF-1, with an optimal dosage for chemotaxis between 10 and 100 ng/ml. After scaffold implantation, the SDF-1-treated group demonstrated complete filling of the cartilage defect with mature cartilage tissue, exhibiting strong proteoglycan content, smooth borders, and good incorporation into marginal cartilage. Modified O’Driscoll scores after 8 weeks showed a significant improvement of cartilage repair in the SDF-1 group relative to the empty control group (P < 0.01), with a trend toward improvement when compared with the gelatin-only-scaffold group (P < 0.1). No significant differences in scores were found between the empty defect group and gelatin-only group.ConclusionIn this study, we demonstrated a simple method for improving the quality of cartilage defect repair in a rat model of microfracture. We confirmed the chemotactic properties of SDF-1 on rat MSCs and found an optimized dosage range for chemotaxis between 10 and 100 ng/ml. Furthermore, we demonstrated a strategy to incorporate SDF-1 into gelatin–collagen I scaffolds in vivo at the site of an osteochondral defect. SDF-1-treated defects displayed robust hyaline cartilage resurfacing of the defect with minimal fibrous tissue, in contrast to the empty control group. The results of the in vitro and in vivo studies together suggest that SDF-1-mediated signaling may significantly improve the quality of cartilage regeneration in an osteochondral defect.