Calf blood flow is augmented in habitually aerobically trained versus untrained postmenopausal women in association with favorable modulation of vasculo-metabolic interactions.

INTRODUCTION: Menopause is associated with vascular dysfunction and increased risk of developing metabolic syndrome. Associations between vascular and metabolic health, and interactions with aerobic exercise training, are unknown in postmenopausal women (PMW). METHODS: In habitually aerobically trained PMW (PMWtr; n = 10; 57 ± 1 years; 40 ± 1 mL/kg/min), strain-gauge plethysmography was used to compare resting and peak calf blood flow (CBFr and CBFpk, respectively) and vascular resistance (CVRr; CVRpk) versus untrained PMW (PMWun; n = 13; 56 ± 1 years; 29 ± 1 mL/kg/min) and premenopausal women (PreM; n = 14; 26 ± 1 years; 40 ± 1 mL/kg/min). Vascular measures were taken before and 1 hour after 45 minutes of aerobic exercise (60% V̇ O2peak ), a known nitric oxide stimulus. Blood analyses included low- (LDLc) and high-density lipoprotein cholesterol (HDLc), insulin, and glucose. RESULTS: Pre-exercise, CBFr and CVRr did not differ (p > 0.05) between PMW groups, nor between PreM and PMWtr. CBFpk was highest (p < 0.05) and CVRpk was lowest (p < 0.05) in PMWtr. Blood markers were similar (p > 0.05) in PMW groups. However, in PMWtr, CBFpk was associated inversely (p < 0.05) with insulin (r = -0.725). Conversely, in PMWun, CBFpk correlated (p < 0.05) inversely with glucose (r = -0.717), positively with HDLc (r = 0.633), and CVRpk positively (p < 0.05) with LDLc (r = 0.568). Post-exercise, CBF increased and CVR decreased (p < 0.05) in all groups, yet CBFpk remained higher and CVRpk lower (p < 0.05) in PMWtr. CONCLUSION: In untrained PMW, peak CBF is associated inversely with circulating pro-atherogenic lipids and glucose. In contrast, peak CBF is associated inversely with insulin levels only in trained PMW. Habitual aerobic exercise may favorably modulate vasculo-metabolic interactions in PMW.

Investigating the Effects of Indirect Coculture of Human Mesenchymal Stem Cells on the Migration of Breast Cancer Cells: A Systematic Review and Meta-Analysis.

Purpose: Breast cancer is the most diagnosed cancer and the leading cause of cancer death in women globally, and mesenchymal stem cells have been widely implicated in tumour progression. This systematic review and meta-analysis seeks to identify and summarise existing literature on the effects of human mesenchymal stem cells (hMSCs) on the migration of breast cancer cells (BCCs) in vitro, to determine the direction of this relationship according to existing research and to identify the directions for future research. Methods: A systematic literature search was conducting using a collection of databases, using the following search terms: in vitro AND mesenchymal stem cells AND breast cancer. Only studies that investigated the effects of human, unmodified MSCs on the migration of human, unmodified BCCs in vitro were included. Standardised mean differences (SMDs) were calculated to determine pooled effect sizes. Results: This meta-analysis demonstrates that hMSCs (different sources combined) increase the migration of both MDA-MB-231 and MCF-7 cell lines in vitro (SMD = 1.84, P = .03 and SMD = 2.69, P < .00001, respectively). Importantly, the individual effects of hMSCs from different sources were also analysed and demonstrated that MSCs derived from human adipose tissue increase BCC migration (SMD = 1.34, P = .0002) and those derived from umbilical cord increased both MDA-MB-231 and MCF-7 migration (SMD = 3.93, P < .00001 and SMD = 3.01, P < .00001, respectively). Conclusions: To our knowledge, this is the first systematic review and meta-analysis investigating and summarising the effects of hMSCs from different sources on the migration of BCCs, in vitro.

Cryopreservation of Human Bone Marrow Derived Mesenchymal Stem Cells at High Concentration Is Feasible.

Introduction: For stem cell therapies to be adopted in mainstream health care, robust, reliable, and cost-effective storage and transport processes must be developed. Cryopreservation remains the best current platform for this purpose, and freezing cells at high concentration may have many benefits, including savings on cost and storage space, facilitating transport logistics, and reducing cryoprotectant volume. Cells, such as mesenchymal stem cells (MSCs), are typically frozen at 1 million cells per milliliter (mL), but the aim of this study is to examine the post-thaw attributes of human bone marrow derived MSCs (hBM-MSCs) frozen at 1, 5, and 10 million cells per mL. Methods: Thawed cells were assessed for their morphology, phenotypic marker expression, viability, apoptosis level, metabolic activity, proliferation, and osteogenic and adipogenic differentiation. Results: In this study, for the first time, it is shown that all assessed cells expressed the typical MSC markers (CD90, CD105, and CD73) and lacked the expression of CD14, CD20, CD34, CD45, and HLA-DR. In addition, all cells showed elongated fibroblastic morphology. Post-thaw viability was retained with no difference among the three concentrations. Moreover, no significant statistical difference was observed in the post-thaw apoptosis level, metabolic activity, proliferation, and osteogenic potential, indicating that these cells are amenable to cryopreservation at higher concentrations. Conclusion: The results of this study are of paramount importance to the development of manufacturing processes around a useful freezing concentration when cells are targeted to be stored for at least 6 months.

The lasting effects of resistance and endurance exercise interventions on breast cancer patient mental wellbeing and physical fitness.

Breast cancer is a persisting global burden for health services with cases and deaths projected to rise in future years. Surgery complemented by adjuvant therapy is commonly used to treat breast cancer, however comes with detrimental side effects to physical fitness and mental wellbeing. The aim of this systematic review and meta-analysis is to determine whether resistance and endurance interventions performed during adjuvant treatment can lastingly ameliorate these side effects. A systematic literature search was performed in various electronic databases. Papers were assessed for bias and grouped based on intervention design. RStudio was used to perform the meta-analyses for each group using the 'meta' package. Publication bias and power analyses were also conducted. These methods conform to PRISMA guidelines. Combined resistance and endurance interventions elicited significant long-lasting improvements in global fatigue and were beneficial to the remaining side effects. Individually, resistance and endurance interventions non-significantly improved these side effects. Resistance interventions elicited higher benefits overall. Exercise interventions have lasting clinical benefits in ameliorating adjuvant therapy side effects, which negatively impact physical fitness and mental wellbeing. These interventions are of clinical value to enhance adherence rates and avoid comorbidities such as sarcopenia, thus improving disease prognosis.

An exploration of the role of exercise in modulating breast cancer progression in vitro: a systematic review and meta-analysis.

Breast cancer is the most prevalent cancer in women worldwide. In the United Kingdom, approximately 5% of all breast cancers are already metastatic at the time of diagnosis. An abundance of literature shows that exercise can have beneficial effects on the outcome and prognosis of breast cancer patients, yet the molecular mechanisms remain poorly understood. There are several in vitro models that aim to recapitulate the response of breast cancer to exercise in vivo; this systematic review and meta-analysis summarizes the existing literature. The following search terms were used to conduct a systematic literature search using a collection of databases (last search performed May 2020): "in vitro," "exercise," and "breast cancer." Only studies that investigated the effects of exercise on breast cancer in vitro were included. Standardized mean differences (SMD) were calculated to determine pooled effect sizes. This meta-analysis has successfully demonstrated that various identified exercise interventions on breast cancer cells in vitro significantly reduced breast cancer cell viability, proliferation, and tumorigenic potential (SMD = -1.76, P = 0.004, SMD = -2.85, P = 0.003, and SMD = -3.15, P = 0.0008, respectively). A clear direction of effect was found with exercise on breast cancer cell migration in vitro, however this effect was not significant (SMD = -0.62, P = 0.317). To our knowledge, this is the first meta-analysis and systematic review investigating and summarizing literature on exercise and breast cancer in vitro, highlighting models used and priority areas for future research focus.

Quantitative assessment of the impact of cryopreservation on human bone marrow-derived mesenchymal stem cells: up to 24 h post-thaw and beyond.

BACKGROUND: The effects of cryopreservation on human bone marrow-derived mesenchymal stem cells (hBM-MSCs) are still ill-defined. In this study, a quantitative approach was adopted to measure several post-thaw cell attributes in order to provide an accurate reflection of the freezing and thawing impact. METHODS: Fresh and cryopreserved passage-matched cells from three different donors were discretely analysed and compared for their viability, apoptosis level, phenotypic marker expression, metabolic activity, adhesion potential, proliferation rate, colony-forming unit ability (CFUF) and differentiation potentials. RESULTS: The results of this study show that cryopreservation reduces cell viability, increases apoptosis level and impairs hBM-MSC metabolic activity and adhesion potential in the first 4 h after thawing. At 24 h post-thaw, cell viability recovered, and apoptosis level dropped but metabolic activity and adhesion potential remained lower than fresh cells. This suggests that a 24-h period is not enough for a full recovery. Beyond 24 h post-thaw, the observed effects are variable for the three cell lines. While no difference is observed in the pre- and post-cryopreservation proliferation rate, cryopreservation reduced the CFUF ability of two of the cell lines and variably affected the adipogenic and osteogenic differentiation potentials of the three cell lines. CONCLUSION: The data collected in this study clearly show that fresh and cryopreserved hBM-MSCs are different, and these differences will inevitably introduce variabilities to the product and process development and subsequently imply financial losses. In order to avoid product divergence pre- and post-cryopreservation, effective strategies to mitigate freezing effects must be developed and implemented.

Characterising hyperinsulinemia-induced insulin resistance in human skeletal muscle cells.

Hyperinsulinaemia potentially contributes to insulin resistance in metabolic tissues, such as skeletal muscle. The purpose of these experiments was to characterise glucose uptake, insulin signalling and relevant gene expression in primary human skeletal muscle-derived cells (HMDCs), in response to prolonged insulin exposure (PIE) as a model of hyperinsulinaemia-induced insulin resistance. Differentiated HMDCs from healthy human donors were cultured with or without insulin (100 nM) for 3 days followed by an acute insulin stimulation. HMDCs exposed to PIE were characterised by impaired insulin-stimulated glucose uptake, blunted IRS-1 phosphorylation (Tyr612) and Akt (Ser473) phosphorylation in response to an acute insulin stimulation. Glucose transporter 1 (GLUT1), but not GLUT4, mRNA and protein increased following PIE. The mRNA expression of metabolic (PDK4) and inflammatory markers (TNF-α) was reduced by PIE but did not change lipid (SREBP1 and CD36) or mitochondrial (UCP3) markers. These experiments provide further characterisation of the effects of PIE as a model of hyperinsulinaemia-induced insulin resistance in HMDCs.

The impact of cryopreservation on bone marrow-derived mesenchymal stem cells: a systematic review.

Mesenchymal stem cells (MSCs) represent an invaluable asset for the field of cell therapy. Human Bone marrow-derived MSCs (hBM-MSCs) are one of the most commonly used cell types in clinical trials. They are currently being studied and tested for the treatment of a wide range of diseases and conditions. The future availability of MSCs therapies to the public will require a robust and reliable delivery process. Cryopreservation represents the gold standard in cell storage and transportation, but its effect on BM-MSCs is still not well established. A systematic review was conducted to evaluate the impact of cryopreservation on BM-MSCs and to attempt to uncover the reasons behind some of the controversial results reported in the literature. Forty-one in vitro studies were analysed, and their results organised according to the cell attributes they assess. It was concluded that cryopreservation does not affect BM-MSCs morphology, surface marker expression, differentiation or proliferation potential. However, mixed results exist regarding the effect on colony forming ability and the effects on viability, attachment and migration, genomic stability and paracrine function are undefined mainly due to the huge variabilities governing the cryopreservation process as a whole and to the lack of standardised assays.

The Role of Dissolved Oxygen Levels on Human Mesenchymal Stem Cell Culture Success, Regulatory Compliance, and Therapeutic Potential.

Most cells in the human body, including human mesenchymal stem cells (hMSCs), have evolved to survive and function in a low physiological oxygen (O2) environment. Investigators have become increasingly aware of the effects of O2 levels on hMSC biology and culture and are mimicking the natural niche of these cells in vitro to improve cell culture yields. This presents many challenges in relation to hMSC identity and function and in the maintenance of a controlled O2 environment for cell culture. The aim of this review was to discuss an "hMSC checklist" as a guide to establishing which identity and potency assays to implement when studying hMSCs. The checklist includes markers, differentiation potential, proliferation and growth, attachment and migration, genomic stability, and paracrine activity. Evidence drawn from the current literature demonstrates that low O2 environments could improve most "hMSC checklist" attributes. However, there are substantial inconsistencies around both the terminology and the equipment used in low O2 studies. Therefore, "hypoxia" as a term and as a culture condition is discussed. The biology of short-term (acute) versus long-term (chronic) hypoxia is considered, and a nascent hypothesis to explain the behavior of hMSCs in long-term hypoxia is presented. It is hoped that by establishing an ongoing discourse and driving toward a regulatory recognizable "hMSC checklist," we may be better able to provide the patient population with safe and efficacious regenerative treatments.

The effect of chronic high insulin exposure upon metabolic and myogenic markers in C2C12 skeletal muscle cells and myotubes.

Skeletal muscle is an insulin sensitive tissue and accounts for approximately 80% of post-prandial glucose disposal. This study describes the effects of insulin, delivered for 72 h, to skeletal muscle myoblasts during differentiation or to skeletal muscle myotubes. After chronic treatment, cultures were acutely stimulated with insulin and analyzed for total and phosphorylated Akt (Ser473 ), mRNA expression of metabolic and myogenic markers and insulin-stimulated glucose uptake. Skeletal muscle cells differentiated in the presence of insulin chronically, reduced acute insulin stimulated phosphorylation of Akt Ser473 . In addition, there was a reduction in mRNA expression of Hexokinase II (HKII), GLUT4 and PGC-1α. Insulin-stimulated glucose uptake was attenuated when cells were differentiated in the presence of insulin. In contrast, myotubes exposed to chronic insulin showed no alterations in phosphorylation of Akt Ser473 . Both HKII and GLUT4 mRNA expression were reduced by chronic exposure to insulin; while PGC-1α was not different between culture conditions and was increased by acute insulin stimulation. These data suggest that there are differential responses in insulin signalling, transcription, and glucose uptake of skeletal muscle cells when cultured in either the presence of insulin during differentiation or in myotube cultures.

Genetic association of pro-inflammatory cytokine gene polymorphisms with coronary artery disease (CAD) in a North Indian population.

BACKGROUND: Cytokines regulate the expression of inflammatory molecules which destabilize the atheromatic plaques. This study focuses on studying the association of inflammatory cytokine polymorphisms like TNF-α -308 (G/A), TNF-ß +252 (A/G), IL-6 -174 (G/C) and IL-6 -597 (G/A), and IFN-É£ +874 (T/A) with coronary artery disease (CAD) among north Indian patients. MATERIALS AND METHODS: 143 CAD and 137 normal healthy controls were recruited in this study. DNA extraction was carried out by high salting out method. TNF-α -308 (G/A) (rs1800797), TNF-ß +252 (A/G) (rs909253), IL-6 -174 (G/C) (rs1800795), IL6 -597 (G/A) (rs1800797), and IFN-É£ +874 (T/A) (rs2430561) SNPs were genotyped by TaqMan®SNP genotyping assays. Different statistical analyses were performed using SPSS v 22.0 and SNPStats. p≤0.05 was considered significant. RESULTS: Significant risk association with CAD was found for TNF-α -308 (G/A) "A" allele (OR=5.6, CI 1.8-17.4, p=0.001) and TNF-ß +252 (A/G) "G" allele (OR=3.4, CI=1.9-6.0, p<0.001). However, no statistical significance was found for IL-6 -174 (G/C) or IL6 -597 (G/A), with CAD. TNF-α -308 (G/A), and TNF-ß +252 (A/G) haplotype "GG" "AG" increased CAD risk significantly (GG haplotype, adjusted OR=2.6, CI 1.4-5.0, p=0.003 and AG haplotype OR=8.5, CI 2.2-33.35, p=0.002) after adjustments for age, sex, TC, TG, HDL, APOB, smoking and diet. DISCUSSION: The present study found significant risk association for TNF-α -308 (G/A), and TNF-ß +252 (A/G) genotypes, alleles and haplotypes, with CAD in a North Indian population.

The Role of TLR4, TNF-α and IL-1ß in Type 2 Diabetes Mellitus Development within a North Indian Population.

This study investigated the role of IL-1ß-511 (rs16944), TLR4-896 (rs4986790) and TNF-α-308 (rs1800629) polymorphisms in type 2 diabetes mellitus (T2DM) among an endogamous Northern Indian population. Four hundred fourteen participants (204 T2DM patients and 210 nondiabetic controls) were genotyped for IL-1ß-511, TLR4-896 and TNF-α-308 loci. The C allele of IL-1ß-511 was shown to increase T2DM susceptibility by 75% (OR: 1.75 [CI 1.32-2.33]). Having two parents affected by T2DM increased susceptibility by 5.7 times (OR: 5.693 [CI 1.431-22.648]). In this study, we have demonstrated a conclusive association with IL-1ß-511 locus and IL-1ß-511-TLR4-896 diplotype (CC-AA) and T2DM, which warrants further comprehensive analyses in larger cohorts.