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Combination treatment of Low-Intensity Vibration (LIV) with zoledronic acid (ZA) was hypothesized to preserve bone mass and muscle strength while reducing adipose tissue accrual associated with complete estrogen (E2)-deprivation in young and skeletally mature mice. Complete E2-deprivation (surgical-ovariectomy (OVX) and daily injection of aromatase inhibitor (AI) letrozole) were performed on 8-week-old C57BL/6 female mice for 4 weeks following commencement of LIV administration or control (no LIV), for 28 weeks. Additionally, 16-week-old C57BL/6 female E2-deprived mice were administered ±LIV twice daily and supplemented with ±ZA (2.5 ng/kg/week). By week 28, lean tissue mass quantified by dual-energy X-ray absorptiometry was increased in younger OVX/AI+LIV(y) mice, with increased myofiber cross-sectional area of quadratus femorii. Grip strength was greater in OVX/AI+LIV(y) mice than OVX/AI(y) mice. Fat mass remained lower in OVX/AI+LIV(y) mice throughout the experiment compared with OVX/AI(y) mice. OVX/AI+LIV(y) mice exhibited increased glucose tolerance and reduced leptin and free fatty acids than OVX/AI(y) mice. Trabecular bone volume fraction and connectivity density increased in the vertebrae of OVX/AI+LIV(y) mice compared to OVX/AI(y) mice; however, this effect was attenuated in the older cohort of E2-deprived mice, specifically in OVX/AI+ZA mice, requiring combined LIV with ZA to increase trabecular bone volume and strength. Similar improvements in cortical bone thickness and cross-sectional area of the femoral mid-diaphysis were observed in OVX/AI+LIV+ZA mice, resulting in greater fracture resistance. Our findings demonstrate that the combination of mechanical signals in the form of LIV and anti-resorptive therapy via ZA improve vertebral trabecular bone and femoral cortical bone, increase lean mass, and reduce adiposity in mice undergoing complete E2-deprivation.One Sentence Summary:Low-magnitude mechanical signals with zoledronic acid suppressed bone and muscle loss and adiposity in mice undergoing complete estrogen deprivation.Translational RelevancePostmenopausal patients with estrogen receptor-positive breast cancer treated with aromatase inhibitors to reduce tumor progression experience deleterious effects to bone and muscle subsequently develop muscle weakness, bone fragility, and adipose tissue accrual. Bisphosphonates (i.e., zoledronic acid) prescribed to inhibit osteoclast-mediated bone resorption are effective in preventing bone loss but may not address the non-skeletal effects of muscle weakness and fat accumulation that contribute to patient morbidity. Mechanical signals, typically delivered to the musculoskeletal system during exercise/physical activity, are integral for maintaining bone and muscle health; however, patients undergoing treatments for breast cancer often experience decreased physical activity which further accelerates musculoskeletal degeneration. Low-magnitude mechanical signals, in the form of low-intensity vibrations, generate dynamic loading forces similar to those derived from skeletal muscle contractility. As an adjuvant to existing treatment strategies, low-intensity vibrations may preserve or rescue diminished bone and muscle degraded by breast cancer treatment.

Erythropoietin (EPO) regulates erythropoiesis by binding to erythropoietin receptor (Epor) on erythroid progenitor cells. Epor is also expressed on bone forming osteoblasts and bone loss accompanies EPO-stimulated erythropoiesis in mice. Mice with Epor restricted to erythroid tissue exhibit reduced bone and increased marrow adipocytes; in contrast, transgenic mice (Tg) with osteoblastic-specific deletion of Epor exhibit reduced trabecular bone with age without change in marrow adipocytes. By 12 weeks, male Tg mice had 22.2% and female Tg mice had 29.6% reduced trabecular bone volume (BV) compared to controls. EPO administration (1200 U/kg) for 10 days reduced trabecular bone in control mice but not in Tg mice. There were no differences in numbers of osteoblasts, osteoclasts, and marrow adipocytes in Tg mice, suggesting independence of EPO signaling in mature osteoblasts, osteoclasts, and adipocytes. Female Tg mice had increased number of dying osteocytes and male Tg mice had a trend for more empty lacunae. Osteogenic cultures from Tg mice had reduced differentiation and mineralization with reduced Alpl and Runx2 transcripts. In conclusion, endogenous EPO-Epor signaling in osteoblasts is important in bone remodeling, particularly trabecular bone and endogenous Epor expression in osteoblasts is required for bone loss accompanying EPO-stimulated erythropoiesis.

Erythropoietin (EPO) is the primary regulator of bone marrow erythropoiesis. Mouse models have provided evidence that EPO also promotes bone remodeling and that EPO‐stimulated erythropoiesis is accompanied by bone loss independent of increased red blood cell production. EPO has been used clinically for three decades to treat anemia in end‐stage renal disease, and notably, although the incidence of hip fractures decreased in the United States generally after 1990, it rose among hemodialysis patients coincident with the introduction and subsequent dose escalation of EPO treatment. Given this clinical paradox and findings from studies in mice that elevated EPO affects bone health, we examined EPO treatment as a risk factor for fractures in hemodialysis patients. Relationships between EPO treatment and hip fractures were analyzed using United States Renal Data System (USRDS) datasets from 1997 to 2013 and Consolidated Renal Operations in a Web‐enabled Network (CROWNWeb) datasets for 2013. Fracture risks for patients treated with

Erythropoietin is essential for bone marrow erythropoiesis and erythropoietin receptor on non-erythroid cells including bone marrow stromal cells suggests systemic effects of erythropoietin. Tg6 mice with chronic erythropoietin overexpression have a high hematocrit, reduced trabecular and cortical bone and bone marrow adipocytes, and decreased bone morphogenic protein 2 driven ectopic bone and adipocyte formation. Erythropoietin treatment (1 200 IU·kg–1) for 10 days similarly exhibit increased hematocrit, reduced bone and bone marrow adipocytes without increased osteoclasts, and reduced bone morphogenic protein signaling in the bone marrow. Interestingly, endogenous erythropoietin is required for normal differentiation of bone marrow stromal cells to osteoblasts and bone marrow adipocytes. ΔEpoRE mice with erythroid restricted erythropoietin receptor exhibit reduced trabecular bone, increased bone marrow adipocytes, and decreased bone morphogenic protein 2 ectopic bone formation. Erythropoietin treated ΔEpoRE mice achieved hematocrit similar to wild-type mice without reduced bone, suggesting that bone reduction with erythropoietin treatment is associated with non-erythropoietic erythropoietin response. Bone marrow stromal cells from wild-type, Tg6, and ΔEpoRE-mice were transplanted into immunodeficient mice to assess development into a bone/marrow organ. Like endogenous bone formation, Tg6 bone marrow cells exhibited reduced differentiation to bone and adipocytes indicating that high erythropoietin inhibits osteogenesis and adipogenesis, while ΔEpoRE bone marrow cells formed ectopic bones with reduced trabecular regions and increased adipocytes, indicating that loss of erythropoietin signaling favors adipogenesis at the expense of osteogenesis. In summary, endogenous erythropoietin signaling regulates bone marrow stromal cell fate and aberrant erythropoietin levels result in their impaired differentiation.

Background: Vaginal candidial infections are due to excessive growth of Candida. These are normally present in the vagina in small numbers. Vaginal infections are typically caused by the yeast species Candida albicans. It is found that candidial infection increases the risk of preterm labour. Aims and objectives of the study were to determine the prevalence of vulvovaginal candidiasis and influence of maternal age, parity and weeks of 2nd trimester on its occurrence among pregnant women in 2nd trimester, attending the antenatal clinic in our hospital.Methods: A prospective study conducted in BMCRI for a period of 3 months (October 2019-December 2019) on patients in second trimester. Consent of patients taken. High vaginal swabs were collected from the pregnant patients in second trimester and sent for culture. Candida positive cases were noted and results were analysed.Results: A total of 100 high vaginal swabs were collected and reported in our study. Among them 54 swabs were positive for Candida growth (54%) and 46 swabs were negative for growth (46%). Culture positive patients’ clinical details were analysed and tabulated.Conclusions: Our study concluded that candidiasis is more prevalent in pregnant women but there was no statistical significance in occurrence of vaginal candidiasis among various age groups, parity or trimester. Hence it is better to screen all the patients in I early II trimester in order to find out and treat positive cases early to prevent preterm births attributed to vaginal candidiasis.

Background: Urinary tract infection (UTI) is one of the many etiological factors of preterm labour. Preterm labour is the onset of labour after 28 weeks and before 37 weeks of gestation. Preterm labour is a significant cause for perinatal morbidity and mortality. Hence early diagnosis and management of etiological factors is necessary. The most common bacterial infection encountered during pregnancy is UTIs. Early detection and management of UTIs may effectively prevent complications of preterm labour including preterm birth. Aims and objectives of the study was to estimate the prevalence of UTIs in preterm labour.Methods: cross sectional study with a total of 250 patients carried out in the department of obstetrics and gynaecology, Vanivilas hospital, BMCRI for a period of 6 moths-Aug 2019-Feb 2020. Patients in preterm labour i.e., cervical dilatation ≥1 cm, cervical effacement ≥80% with true labour pains were included in the study after taken informed consent form the patient. Detailed clinical history including age of patient, level of education, duration of antenatal care, parity, and obstetrical history was taken. Gestational age was calculated by LMP or early ultrasound. General examination, systemic examination and obstetric examination was done. Routine investigations were done along with clean catch midstream urine sample in a sterile container. Two samples were collected: 1st sample for microscopic examination, 2nd sample for culture and sensitivity.Conclusions: Untreated UTI can be associated with obstetric complications. The most common bacterial infection during pregnancy is UTIs. All women should be screened for UTI at the first antenatal visit. Once diagnosed it should be promptly treated with suitable antibiotic which is sensitive yet safest.

Obesity remodels bone and increases bone marrow adipocytes (BMAT), which negatively regulate hematopoiesis and bone. Reduced BMAT could restore altered hematopoiesis and bone features. We analyzed the potential of erythropoietin (EPO), the cytokine required for erythropoiesis, to inhibit BMAT in C57BL6/J mice fed four weeks of a high-fat diet (HFD). Acute EPO administration markedly decreased BMAT in regular chow diet (RCD) and HFD-fed mice, without affecting whole body fat mass. Micro-CT analysis showed EPO reduced trabecular bone in RCD- and HFD-fed mice, but EPO-treated HFD-fed mice maintained cortical bone mineral density and cortical bone volume, which was reduced on RCD. Despite achieving similar increased hematocrits with BMAT loss in RCD- and HFD-fed mice treated with EPO, decreased bone marrow cellularity was only observed in RCD-fed mice concomitant with an increasing percentage of bone marrow erythroid cells. In contrast, in HFD-fed mice, EPO increased endothelial cells and stromal progenitors with a trend toward the normalization of marrow homeostasis. EPO administration increased c-terminal FGF23 and intact serum FGF23 only in HFD-fed mice. These data demonstrate the distinct EPO responses of bone and marrow in normal and obese states, accompanying EPO-induced loss of BMAT.

Background: The development of doppler ultrasonographic technology has provided an opportunity to obtain a qualitative and quantitative assessment of maternal and foetal circulation using a non-invasive method. It has been proved by many studies that doppler has a very important role in screening of high-risk pregnancies. Objective of this study was to evaluate the role of colour doppler study in normal and high-risk pregnancy in relation to perinatal outcome.Methods: A prospective study was done including 75 women with high risk pregnancy and 75 normal pregnant women during the period October 2018 to September 2019 in hospitals attached to Bangalore Medical College and Research Institute. Doppler examination was done after recording patients’ history, clinical examination and ultrasound. Results were analysed and conclusions were made.Results: Out of the 22 patients with PIH, 20 patients had abnormal umbilical artery S/D ratio and all 22 had abnormal MCA PI. Out of 12 patients with diabetes, 10 had abnormal umbilical artery S/D ratio. All the patients with IUGR had abnormal umbilical artery S/D ratio and abnormal MCA PI.Conclusions: Colour doppler flow velocimetry done repeatedly can predict adverse foetal events with a great degree of accuracy.

Erythropoietin (EPO) regulation of red blood cell production and its induction at reduced oxygen tension provides for the important erythropoietic response to ischemic stress. The cloning and production of recombinant human EPO has led to its clinical use in patients with anemia for two and half decades and has facilitated studies of EPO action. Reports of animal and cell models of ischemic stress in vitro and injury suggest potential EPO benefit beyond red blood cell production including vascular endothelial response to increase nitric oxide production, which facilitates oxygen delivery to brain, heart and other non-hematopoietic tissues. This review discusses these and other reports of EPO action beyond red blood cell production, including EPO response affecting metabolism and obesity in animal models. Observations of EPO activity in cell and animal model systems, including mice with tissue specific deletion of EPO receptor (EpoR), suggest the potential for EPO response in metabolism and disease.

Background: Bleeding in first and second trimester of pregnancy is one of the common complications of pregnancy. there is evidence from various prospective and retrospective studies that first and second trimester vaginal bleeding which continue with pregnancy is associated with adverse pregnancy outcome, including preterm delivery, low birth weight babies, perinatal death and congenital anomalies. Objective of this study was to know the outcome of pregnancies who have bleeding in first and second trimester of pregnancy.Methods: This study was prospective study done in the department of obstetrics and gynaecology, Vanivilas Hospital, Bangalore from September 2018 to August 2019.Results: This study concludes that I trimester vaginal bleeding are at increased risk of abortion than in II trimester vaginal bleeding. Risk of placenta previa was more in II trimester vaginal bleeding than in I trimester vaginal bleeding.Conclusions: This study concludes that I trimester vaginal bleeding are at increased risk of abortion than in II trimester vaginal bleeding. Risk of placenta previa was more in II trimester vaginal bleeding than in I trimester vaginal bleeding. Bleeding in I trimester and II trimester call for special attention in view of increased risk of preterm birth and perinatal death. Recognition of these association will be useful for detection and follow up of pregnancies being at high risk.

Deregulated NOTCH1 has been reported in lymphoid leukaemia, although its role in chronic myeloid leukaemia (CML) is not well established. We previously reported BCR-ABL down-regulation of a novel haematopoietic regulator, CCN3, in CML; CCN3 is a non-canonical NOTCH1 ligand. This study characterizes the NOTCH1–CCN3 signalling axis in CML. In K562 cells, BCR-ABL silencing reduced full-length NOTCH1 (NOTCH1-FL) and inhibited the cleavage of NOTCH1 intracellular domain (NOTCH1-ICD), resulting in decreased expression of the NOTCH1 targets c-MYC and HES1. K562 cells stably overexpressing CCN3 (K562/CCN3) or treated with recombinant CCN3 (rCCN3) showed a significant reduction in NOTCH1 signalling (> 50% reduction in NOTCH1-ICD, p < 0.05). Gamma secretase inhibitor (GSI), which blocks NOTCH1 signalling, reduced K562/CCN3 colony formation but increased that of K562/control cells. GSI combined with either rCCN3 or imatinib reduced K562 colony formation with enhanced reduction of NOTCH1 signalling observed with combination treatments. We demonstrate an oncogenic role for NOTCH1 in CML and suggest that BCR-ABL disruption of NOTCH1–CCN3 signalling contributes to the pathogenesis of CML.

Chronic Myeloid Leukaemia (CML) is a myeloproliferative disorder characterized by the expression of the oncoprotein, Bcr-Abl kinase. CCN3 normally functions as a negative growth regulator, but it is downregulated in CML, the mechanism of which is not known. MicroRNAs (miRNAs) are small non-coding RNAs, which negatively regulate protein translation by binding to the complimentary sequences of the 3′ UTR of messenger RNAs. Deregulated miRNA expression has emerged as a hallmark of cancer. In CML, BCR-ABL upregulates oncogenic miRNAs and downregulates tumour suppressor miRNAs favouring leukaemic transformation. We report here that the downregulation of CCN3 in CML is mediated by BCR-ABL dependent miRNAs. Using the CML cell line K562, we profiled miRNAs, which are BCR-ABL dependent by transfecting K562 cells with anti-BCR-ABL siRNA. MiRNA expression levels were quantified using the Taqman Low Density miRNA array platform. From the miRNA target prediction databases we identified miRNAs that could potentially bind to CCN3 mRNA and reduce expression. Of these, miR-130a, miR-130b, miR-148a, miR-212 and miR-425-5p were significantly reduced on BCR-ABL knockdown, with both miR-130a and miR-130b decreasing the most within 24 h of siRNA treatment. Transfection of mature sequences of miR-130a and miR-130b individually into BCR-ABL negative HL60 cells resulted in a decrease of both CCN3 mRNA and protein. The reduction in CCN3 was greatest with overexpression of miR-130a whereas miR-130b overexpression resulted only in marginal repression of CCN3. This study shows that miRNAs modulate CCN3 expression. Deregulated miRNA expression initiated by BCR-ABL may be one mechanism of downregulating CCN3 whereby leukaemic cells evade negative growth regulation.

Erythropoietin receptor (EPOR) expression level determines the extent of erythropoietin (EPO) response. Previously we showed that EPOR expression in endothelial cells is increased at low oxygen tension and that EPO stimulation of endothelial cells during hypoxia can increase endothelial nitric oxide (NO) synthase (eNOS) expression and activation as well as NO production. We now observe that while EPO can stimulate NO production, NO in turn can regulate EPOR expression. Human umbilical vein endothelial cells (HUVEC) treated with 10–50 μM of NO donor diethylenetriamine NONOate (DETANO) for 24 hours showed significant induction of EPOR gene expression at 5% and 2% of oxygen. Also human bone marrow microvascular endothelial cell line (TrHBMEC) cultured at 21 and 2% oxygen with 50 μM DETANO demonstrated a time and oxygen dependent induction of EPOR mRNA expression after 24 and 48 hours, particularly at low oxygen tension. EPOR protein was also induced by DETANO at 2% oxygen in TrHBMEC and HUVEC. The activation of signaling pathways by NO donor stimulation appeared to be distinct from EPO stimulation. In reporter gene assays, DETANO treatment of HeLa cells at 2% oxygen increased EPOR promoter activity indicated by a 48% increase in luciferase activity with a 2 kb EPOR promoter fragment and a 71% increase in activity with a minimal EPOR promoter fragment containing 0.2Kb 5′. We found that DETANO activated MAPK kinase in TrHBMEC both in normoxia and hypoxia, while MAPK kinase inhibition showed significant reduction of EPOR mRNA gene expression at low oxygen tension, suggesting MAPK involvement in NO mediated induction of EPOR. Furthermore, DETANO stimulated Akt anti-apoptotic activity after 30 minutes in normoxia, whereas it inhibited Akt phosphorylation in hypoxia. In contrast, EPO did not significantly increase MAPK activity while EPO stimulated Akt phosphorylation in TrHBMEC in normoxia and hypoxia. These observations provide a new effect of NO on EPOR expression to enhance EPO response in endothelial cells, particularly at low oxygen tensions.

The NOTCH pathway is an evolutionarily conserved signalling network, which is fundamental in regulating developmental processes in invertebrates and vertebrates (Gazave et al. in BMC Evol Biol 9:249, 2009). It regulates self-renewal (Butler et al. in Cell Stem Cell 6:251–264, 2010), differentiation (Auderset et al. in Curr Top Microbiol Immunol 360:115–134, 2012), proliferation (VanDussen et al. in Development 139:488–497, 2012) and apoptosis (Cao et al. in APMIS 120:441–450, 2012) of diverse cell types at various stages of their development. NOTCH signalling governs cell-cell interactions and the outcome of such responses is highly context specific. This makes it impossible to generalize about NOTCH functions as it stimulates survival and differentiation of certain cell types, whereas inhibiting these processes in others (Meier-Stiegen et al. in PLoS One 5:e11481, 2010). NOTCH was first identified in 1914 in Drosophila and was named after the indentations (notches) present in the wings of the mutant flies (Bigas et al. in Int J Dev Biol 54:1175–1188, 2010). Homologs of NOTCH in vertebrates were initially identified in Xenopus (Coffman et al. in Science 249:1438–1441, 1990) and in humans NOTCH was first identified in T-Acute Lymphoblastic Leukaemia (T-ALL) (Ellisen et al. in Cell 66:649–61, 1991). NOTCH signalling is integral in neurogenesis (Mead and Yutzey in Dev Dyn 241:376–389, 2012), myogenesis (Schuster-Gossler et al. in Proc Natl Acad Sci U S A 104:537–542, 2007), haematopoiesis (Bigas et al. in Int J Dev Biol 54:1175–1188, 2010), oogenesis (Xu and Gridley in Genet Res Int 2012:648207, 2012), differentiation of intestinal cells (Okamoto et al. in Am J Physiol Gastrointest Liver Physiol 296:G23–35, 2009) and pancreatic cells (Apelqvist et al. in Nature 400:877–881, 1999). The current review will focus on NOTCH signalling in normal and malignant blood cell production or haematopoiesis.