A NOTCH3-CXCL12-driven myeloma-tumor niche signaling axis promotes chemoresistance in multiple myeloma.

Multiple myeloma (MM) remains incurable due to disease relapse and drug resistance. Notch signals from the tumor microenvironment (TME) confer chemoresistance, but the cellular and molecular mechanisms are not entirely understood. Using clinical and transcriptomic datasets, we found that NOTCH3 is upregulated in CD138+ cells from newly diagnosed MM (NDMM) patients compared to healthy individuals and increased in progression/relapsed MM (PRMM) patients. Further, NDMM patients with high NOTCH3 expression exhibited worse responses to bortezomib (BOR)-based therapies. Cells of the TME, including osteocytes, upregulated NOTCH3 in MM cells and protected them from apoptosis induced by BOR. NOTCH3 activation (NOTCH3OE) in MM cells decreased BOR anti-MM efficacy and its ability to improve survival in in vivo myeloma models. Molecular analyses revealed that NDMM and PRMM patients with high NOTCH3 exhibit CXCL12 upregulation. TME cells upregulated CXCL12 and activated the CXCR4 pathway in MM cells in a NOTCH3-dependent manner. Moreover, genetic or pharmacologic inhibition of CXCL12 in NOTCH3OE MM cells restored sensitivity to BOR regimes in vitro and in human bones bearing NOTCH3OE MM tumors cultured ex vivo. Our clinical and preclinical data unravel a novel NOTCH3-CXCL12 pro-survival signaling axis in the TME and suggest that osteocytes transmit chemoresistance signals to MM cells.

Fenofibrate attenuates ischemia reperfusion-induced acute kidney injury and associated liver dysfunction in rats.

Ischemia/reperfusion (I/R) is one of the common reasons for acute kidney injury (AKI) and we need to develop effective therapies for treating AKI. We investigated the role of fenofibrate against I/R-induced AKI and associated hepatic dysfunction in rats. In male wistar albino rats, renal pedicle occlusion for 40 min and 24 h reperfusion resulted in AKI. I/R-induced AKI was demonstrated by measuring serum creatinine, creatinine clearance, urea, uric acid, potassium, fractional excretion of sodium and urinary microproteins. Oxidative stress in rat kidneys was quantified by assaying superoxide anion generation, thiobarbituric acid reactive substances, and reduced glutathione levels. AKI-induced hepatic damage was quantified by assaying serum aminotransferases, alkaline phosphatase and bilirubin levels. Moreover, serum cholesterol, high density lipoprotein and triglycerides were quantified. Hematoxylin-eosin staining of renal and hepatic tissues was done and the kidney and liver injury scores were determined. Immunohistology of endothelial nitric oxide synthase (eNOS) was done in rat kidneys. Fenofibrate was administered for 1 week before subjecting rats to AKI. In separate group, the nitric oxide synthase inhibitor, L-nitroarginine methyl ester (L-NAME) was administered prior to fenofibrate treatment. In I/R group, significant alteration in the serum/urine parameters indicated AKI and hepatic dysfunction along with marked increase in kidney and liver injury scores. Treatment with fenofibrate attenuated AKI and associated hepatic dysfunction. Moreover, I/R-induced decrease in renal eNOS expression was abrogated by fenofibrate. Pre-treatment with L-NAME abolished fenofibrate mediated reno- and hepato-protective effects. In conclusion, fenofibrate attenuates I/R-induced AKI and associated hepatic dysfunction putatively through modulation of eNOS expression.

GSK3ß Inhibition Restores Impaired Neurogenesis in Preterm Neonates With Intraventricular Hemorrhage.

Intraventricular hemorrhage (IVH) is a common complication of prematurity in infants born at 23-28 weeks of gestation. Survivors exhibit impaired growth of the cerebral cortex and neurodevelopmental sequeale, but the underlying mechanism(s) are obscure. Previously, we have shown that neocortical neurogenesis continues until at least 28 gestational weeks. This renders the prematurely born infants vulnerable to impaired neurogenesis. Here, we hypothesized that neurogenesis is impaired by IVH, and that signaling through GSK3ß, a critical intracellular kinase regulated by Wnt and other pathways, mediates this effect. These hypotheses were tested observationally in autopsy specimens from premature infants, and experimentally in a premature rabbit IVH model. Significantly, in premature infants with IVH, the number of neurogenic cortical progenitor cells was reduced compared with infants without IVH, indicating acutely decreased neurogenesis. This finding was corroborated in the rabbit IVH model, which further demonstrated reduction of upper layer cortical neurons after longer survival. Both the acute reduction of neurogenic progenitors, and the subsequent decrease of upper layer neurons, were rescued by treatment with AR-A014418, a specific inhibitor of GSK3ß. Together, these results indicate that IVH impairs late stages of cortical neurogenesis, and suggest that treatment with GSK3ß inhibitors may enhance neurodevelopment in premature infants with IVH.

Osteocyte Cellular Senescence.

PURPOSE OF REVIEW: Senescent cells are now known to accumulate in multiple tissues with aging and through their inflammation (the senescence-associated secretory phenotype, SASP) contribute to aging and chronic diseases. Here, we review the roles of senescent osteocytes in the context of bone loss. RECENT FINDINGS: Numerous studies have established that senescent osteocytes accumulate in the bone microenvironment with aging in mice and in humans. Moreover, at least in mice, elimination of senescent cells results in attenuation of age-related bone loss. Osteocyte senescence also occurs in response to other cellular stressors, including radiotherapy, chemotherapy, and metabolic dysfunction, where it appears to mediate skeletal deterioration. Osteocyte senescence is linked to bone loss associated with aging and other conditions. Senescent osteocytes are potential therapeutic targets to alleviate skeletal dysfunction. Additional studies better defining the underlying mechanisms as well as translating these exciting findings from mouse models to humans are needed.

Test-Retest Reliability Between Free Weight and Machine-Based Movement Velocities.

Miller, RM, Freitas, EDS, Heishman, AD, Koziol, KJ, Galletti, BAR, Kaur, J, and Bemben, MG. Test-retest reliability between free weight and machine-based movement velocities. J Strength Cond Res 34(2): 440-444, 2020-Several devices are available to measure muscular power through velocity measurement, including the Tendo FitroDyne. The ability for such devices to produce consistent results is still questioned, and the reproducibility of measurement between free weight and machine exercise has yet to be examined. Therefore, the aim of this investigation was to determine the test-retest reliability for barbell velocity during the bench press (BP) and weight velocity during the 2 leg press (2LP) for loads corresponding to 20-80% of 1 repetition maximum (1RM). Forty recreationally active individuals (22.6 ± 2.5 years; 175.9 ± 10.8 cm; and 76.2 ± 13.2 kg) with a 1RM BP and 2LP of 66.8 ± 32.4 kg and 189.5 ± 49 kg, respectively, volunteered for this study. Subjects completed 1 familiarization visit preceding 3 testing visits, which encompassed 1RM determination and 2 days of velocity testing. Forty-eight hours after 1RM testing, the subjects performed 1 repetition at 20, 30, 40, 50, 60, 70, and 80% of their 1RM for each exercise in randomized order. Subjects returned to the laboratory 1 week later to perform the velocity assessment again in randomized order. Intraclass correlation coefficient (ICC2,1) and relative SEM for the BP and 2LP ranged from 0.56 to 0.98 (3-18.1%) and 0.78 to 0.98 (2.8-7.2%), respectively, and no mean differences were observed between trials. The results suggest high reliability for BP velocity between 30 and 60% 1RM and moderate reliability at 20, 70, and 80% 1RM, while the 2LP displayed high to excellent reliability from 20 to 80% 1RM. Cumulatively, machine-based exercise displayed greater reproducibility; however, additional machine exercises need to be examined to bolster this conclusion.

Maximal power production as a function of sex and training status.

Maximal muscular power is achieved at lower percentages of maximal strength (1RM); however, this notion has not been elucidated based on sex or training status. Therefore, the purpose of this investigation was to examine the influence of sex and training status on maximal power production. Sixty men and women (resistance trained or untrained) completed 1RM testing for the two-leg press (2LP) and bench press (BP). Participants then returned to perform single repetitions at 20, 30, 40, 50, 60, 70 and 80% of their 1RM to determine muscular power. Factorial analyses determined significant interactions (training status by sex by intensity) for the BP (F=35.6, p<0.001) and 2LP (F=8.2, p<0.001). Subsequent analyses indicated that during the BP trained men produce maximal power between 30-40% 1RM compared to untrained men at 60-70% 1RM. Trained women produced maximal power at 50% 1RM compared to untrained women at 60-70% 1RM. During the 2LP, trained men produced maximal power at 40% 1RM compared to untrained men at 60% 1RM. Trained women produced maximal power at 50% 1RM compared to 60-70% 1RM in untrained women. These data suggest that resistance trained individuals and men display maximal power at a lower relative intensity than untrained individuals and women.

Effect of Aggregation on the Hydrodynamic Properties of Bovine Serum Albumin.

PURPOSE: To systematically analyze shape and size of soluble irreversible aggregates and the effect of aggregate formation on viscosity. METHODS: Online light scattering, refractive index and viscosity detectors attached to HPLC (Viscotek®) were used to study aggregation, molecular weight and intrinsic viscosity of bovine serum albumin (BSA). Irreversible aggregates were generated by heat stress. Bulk viscosity was measured by an oscillating piston viscometer. RESULTS: As BSA was heated at a higher concentration or for a longer time, the relative contribution, molecular weight and intrinsic viscosity of aggregate species increased. Molecular shape was evaluated from intrinsic viscosity values, and aggregates were estimated to be more asymmetric than monomer species. The presence of aggregates resulted in an increase in bulk viscosity when relative contribution of very high molecular weight species exceeded 10%. CONCLUSIONS: For model system and conditions studied, generation of higher order aggregate species was concluded to be associated with an increase in molecular asymmetry. Elevated viscosity in the presence of aggregated species points to molecular asymmetry being a critical parameter affecting solution viscosity of BSA.

Time Course Change in Muscle Swelling: High-Intensity vs. Blood Flow Restriction Exercise.

This study determined the time course for changes in muscle swelling and plasma volume following high (HI) and low-intensity resistance exercise with blood-flow restriction (LI-BFR). Ten male participants (22.1±3.0 yrs) completed three experimental conditions: high-intensity exercise (HI - 80% of 1RM), low-intensity exercise with BFR (LI-BFR -20% of 1RM, and 160 mmHg of BFR), and control (CON - no exercise or BFR). Muscle cross-sectional area (mCSA), muscle thickness, thigh circumference, and percentage change in plasma volume (PV%∆) were measured. mCSA was significantly greater than rest values at 15 min post-exercise (p<0.01) for HI and LI-BFR, and at 75 min post-exercise (p<0.01) for HI. Muscle thickness was significantly greater than rest immediately post-exercise (p<0.01) and 30 min post-exercise (p<0.01) for HI and LI-BFR, and at 60 min post-exercise for HI (p=0.01). Muscle thickness was greater for BFR immediately post-exercise compared to HI (p=0.01) post-exercise. Thigh circumference was significantly greater from rest at 15 min post-exercise (p=0.01) and at 75 min post-exercise for both LI-BFR (p=0.03) and HI (p<0.01). PV%∆ significantly decreased from rest immediately post-exercise for both HI (p<0.01) and LI-BFR (p<0.01). In conclusion, BFR exercise induces changes in muscle swelling and plasma volume similar to those observed at high-intensities.

Management of Benign Paroxysmal Positional Vertigo: A Comparative Study between Epleys Manouvre and Betahistine.

ABSTRACT: Benign paroxysmal positional vertigo (BPPV) is the most common peripheral vestibular disorder, accounting for 20% of all vertigo cases. Idiopathic BPPV is most common between the ages of 50 and 70, although the condition is found in all age groups. This study was conducted in our institute on 90 patients who presented to the outpatient department with history of vertigo and were diagnosed with BPPV via a positive Dix Hallpike test. Patients were randomnly placed in three groups of 30 each. Patients in Group A were treated with Epleys manoeuvre alone, in Group B were treated with Epleys Manouvre followed by oral Betahistine and patients in Group C were treated with Betahistine alone. All the patients were followed up after 1 week and 4 weeks following treatment. In our study we found that patients responded better when they were treated with Epleys Manouvre with Betahistine with less relapse and recurrence. Treatment with Epleys manouvre resulted in early improvement of symptoms. It was found in our study that Betahistine as a sole modality of treatment of vertigo in BPPV can be preferred in patients who are unfit to undergo canal repositioning manouvres.

Assessment of demand for male contraceptives: A multi-country study.

BACKGROUND: As of 2019, high unmet contraceptive needs in low- and middle-income countries result in nearly half of all pregnancies being unintended. Additional male contraceptive options could help, but gaps remain in assessing demand for them. As the development of novel male contraceptives progresses, addressing these gaps would help inform policy and scientific direction. OBJECTIVES: Measure consumer demand for novel male contraceptives and identify product attributes associated with men's preferences. MATERIALS & METHODS: A cross-sectional, probability-based-sample survey was conducted with 12,435 sexually active, fertile cisgender, heterosexual men aged 18-60 years, and 9,122 of their female partners in Bangladesh, Vietnam, Kenya, Nigeria, DR Congo, and Côte d'Ivoire from May 2021 to October 2022. The survey was also conducted in the United States among 3,243 of these men in April 2022. A follow-up with 3,070 men in May 2023 assessed potential changes in demand after the Supreme Court's Dobbs v. Jackson Women's Health Organization decision. The surveys assessed contraceptive use, perceptions, and sociodemographic characteristics, and a discrete choice experiment evaluated preferences for 11 potential product attributes. Data from female partners in the six LMICs indicated their interest and trust in male partners' contraceptive use. RESULTS: On average, 61% of men showed interest in trying novel male contraceptives in their first year of availability, ranging from 39% and 49% (pre- and post-Dobbs) in the United States to 76% in Nigeria and Bangladesh. Form of administration and time of use drove men's product preferences. Female partners' interest and trust in male contraceptive use were high across geographies. DISCUSSION: This study reveals strong latent demand for novel male contraceptives, highlighting the potential for novel male methods to meet unmet needs, and female partners' trust in men to use them. CONCLUSION: High demand for novel male contraceptives among men and their partners exists, supporting investment in their development.

Effect of telephone reminders on adherence to anti-rabies vaccine amongst animal bite patients in North India: A randomised, single-blind, parallel-group, single-centre, interventional superiority trial.

Objectives: Rabies is a highly infectious viral zoonotic disease of the central nervous system with a near 100% fatality rate. Vaccine adherence is an integral part of achieving effective treatment. India accounts for 27% of the global deaths from rabies yearly. Rabid dog bites are responsible for 99% of these deaths. This study aimed to assess the effect of reminder calls on compliance with the anti-rabies vaccine among animal bite patients. Methods: An interventional, randomised, single-blinded, parallel-group, single-centre study was conducted at the Anti Rabies Clinic, Rajindra Hospital, Government Medical College, Patiala, a city located in Punjab, India, with a population of approximately 19 lakhs. A sample of 400 patients was enrolled and divided into two groups by lottery method. After obtaining written and informed consent from patients, data were collected using a validated pre-tested, semi-structured proforma. The intervention group received reminder calls before each dose. At the end of the study, complete information regarding compliance was obtained from both groups and analysed. Results: The median age group of the animal bite patients was 21-40 years, with most being male (69.50%). Most of the bites were on the lower extremities (64.0%), followed by the upper extremities (29.0%) and the face (3.25%). Out of 153 patients who delayed the dose, 137 (89.54%) delayed a single dose. The 4th dose on the 28th day was the most frequently delayed dose (75.16%). Reminder calls increased the vaccine compliance rate from 53.5% in the non-intervention group to 70% in the intervention group (adjusted odds ratio=2.28; P=0.0002). There was no effect of gender, area, educational qualification, or marital status on the compliance. Conclusions: Reminder calls were found to have significant effect on the adherence to the anti-rabies vaccine. This simple, cost-effective, and patient-friendly intervention must be integrated within the health care system to ensure timely and complete administration of the anti-rabies vaccine to reduce the risk of rabies.

Lack of documentation in animal bite cases and its impact on rabies biologicals utilization.

Introduction: Rabies has significant health and economic consequences for both humans and animals. Annually, India witnesses 17.4 million dog bites, yet only 3 million individuals receive post-exposure prophylaxis (PEP). There is a shortage of anti-rabies vaccine in India as quoted in many news reports. In India, lack of documentation of previous vaccination against animal bites is there, hence resulting in the re-administration of the anti-rabies vaccine, leading to a significant biological loss (anti-rabies vaccine). Material and Methods: A cross-sectional, retrospective study was conducted. Data was collected, and analyzed from June 2021 to June 2023 a period of 2 years. Results: Majority of the patients reported within the first 24 hours after being bitten while approximately one-third reported after 24 hours. Majority were Category 3 bites and unprovoked. Males, lower-middle class, and bites on lower extremities were common among 4291 patients attending the clinic. Out of 217 re-exposure cases, 185 did not have any documentation regarding their previous treatment of animal bites. Conclusion: Among 4291 patients attending the clinic, majority were Category 3 bites on the lower extremities. 85.25% of re-exposure cases had to be administered a full course of treatment due to a lack of documentation leading to rabies as a biological wastage. This avoidable wastage can be a resource for treating more patients.

Single-cell Transcriptome Analysis Identifies Senescent Osteocytes as Contributors to Bone Destruction in Breast Cancer Metastasis.

Breast cancer bone metastases increase fracture risk and are a major cause of morbidity and mortality among women. Upon colonization by tumor cells, the bone microenvironment undergoes profound reprogramming to support cancer progression that disrupts the balance between osteoclasts and osteoblasts, leading to bone lesions. Whether such reprogramming affects matrix-embedded osteocytes remains poorly understood. Here, we demonstrate that osteocytes in breast cancer bone metastasis develop premature senescence and a distinctive senescence-associated secretory phenotype (SASP) that favors bone destruction. Single-cell RNA sequencing identified osteocytes from mice with breast cancer bone metastasis enriched in senescence and SASP markers and pro-osteoclastogenic genes. Using multiplex in situ hybridization and AI-assisted analysis, we detected osteocytes with senescence-associated distension of satellites, telomere dysfunction, and p16Ink4a expression in mice and patients with breast cancer bone metastasis. In vitro and ex vivo organ cultures showed that breast cancer cells promote osteocyte senescence and enhance their osteoclastogenic potential. Clearance of senescent cells with senolytics suppressed bone resorption and preserved bone mass in mice with breast cancer bone metastasis. These results demonstrate that osteocytes undergo pathological reprogramming by breast cancer cells and identify osteocyte senescence as an initiating event triggering bone destruction in breast cancer metastases.

MicroRNA-19a-3p Decreases with Age in Mice and Humans and Inhibits Osteoblast Senescence.

Aging is a major risk factor for most chronic diseases, including osteoporosis, and is characterized by an accumulation of senescent cells in various tissues. MicroRNAs (miRNAs) are critical regulators of bone aging and cellular senescence. Here, we report that miR-19a-3p decreases with age in bone samples from mice as well as in posterior iliac crest bone biopsies of younger versus older healthy women. miR-19a-3p also decreased in mouse bone marrow stromal cells following induction of senescence using etoposide, H2O2, or serial passaging. To explore the transcriptomic effects of miR-19a-3p, we performed RNA sequencing of mouse calvarial osteoblasts transfected with control or miR-19a-3p mimics and found that miR-19a-3p overexpression significantly altered the expression of various senescence, senescence-associated secretory phenotype-related, and proliferation genes. Specifically, miR-19a-3p overexpression in nonsenescent osteoblasts significantly suppressed p16 Ink4a and p21 Cip1 gene expression and increased their proliferative capacity. Finally, we established a novel senotherapeutic role for this miRNA by treating miR-19a-3p expressing cells with H2O2 to induce senescence. Interestingly, these cells exhibited lower p16 Ink4a and p21 Cip1 expression, increased proliferation-related gene expression, and reduced SA-ß-Gal+ cells. Our results thus establish that miR-19a-3p is a senescence-associated miRNA that decreases with age in mouse and human bones and is a potential senotherapeutic target for age-related bone loss. © 2023 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Predictive factors for nodal recurrence in differentiated thyroid cancers.

INTRODUCTION: Differentiated thyroid carcinoma usually has a good prognosis. Primary treatment is surgery, followed by radioactive iodine ablation based on risk stratification. The incidence of local and distant recurrence is 30%. Recurrence can be managed surgically or with multiple cycles of radioactive iodine ablation. There are multiple risk factors for structural disease recurrence proposed by the American Thyroid Association. In this study, we attempted to study the risk factors of structural recurrence in differentiated carcinoma thyroid and the pattern of recurrence in patients with node negative thyroid cancer who underwent total thyroidectomy. METHODOLOGY: This study selected a retrospective cohort of 1498 patients with differentiated thyroid cancer: out of these, 137 patients who presented after thyroidectomy with cervical nodal recurrence from January 2017 to December 2020 were included. The risk factors for central and lateral lymph node metastasis were analysed by univariate and multivariate analyses, including age, gender, T-stage, extrathyroidal extension, multifocality and high-risk variants. In addition, the presence of TERT/BRAF mutations was studied as a risk factor for central and lateral nodal recurrence. RESULTS: Out of 1498 patients, 137 who fit the inclusion criteria were analysed. Majority were female (73%); mean age was 43.1 years. Lateral compartment neck nodal recurrence was more common (84%), while isolated central compartment nodal recurrence occurred only in 16%. Most recurrences were seen in the first 1 year (23.3%) or after 10 years post-total thyroidectomy (35.7%). On univariate variate analysis, multifocality, extrathyroidal extension and high-risk variants stage were significant factors for nodal recurrence. However, on multivariate analysis for lateral compartment recurrence, multifocality, extrathyroidal extension and age were found to be significant. On multivariate analysis, multifocality, extrathyroidal extension and presence of high-risk variants were significant predictors of central compartment nodal metastasis. ROC curve analysis showed AUC for ETE (AUC-0.795), multifocality (AUC-0.860), presence of high-risk variants (AUC-0.727) and T-stage (AUC-0.771) as sensitive predictive factors for central compartment. 69 percent patients with very early recurrences (<6 month) had TERT/BRAF V600 E mutations. CONCLUSION: In our study, we have noted extrathyroidal extension and multifocality as significant risk factors for nodal recurrence. BRAF and TERT mutations are associated with aggressive clinical course and early recurrences. There is limited role of prophylactic central compartment node dissection.

Multiparametric senescent cell phenotyping reveals CD24 osteolineage cells as targets of senolytic therapy in the aged murine skeleton.

Senescence drives organismal aging, yet the deep characterization of senescent cells in vivo remains incomplete. Here, we applied mass cytometry by time-of-flight (CyTOF) using carefully validated antibodies to analyze senescent cells at single-cell resolution. We used multiple criteria to identify senescent mesenchymal cells that were growth arrested and resistant to apoptosis (p16+/Ki67-/BCL-2+; "p16KB" cells). These cells were highly enriched for senescence-associated secretory phenotype (SASP) and DNA damage markers and were strongly associated with age. p16KB cell percentages were also increased in CD24+ osteolineage cells, which exhibited an inflammatory SASP in aged mice and were robustly cleared by both genetic and pharmacologic senolytic therapies. Following isolation, CD24+ skeletal cells exhibited growth arrest, SA-ßgal positivity, and impaired osteogenesis in vitro . These studies thus provide a new approach using multiplexed protein profiling by CyTOF to define senescent mesenchymal cells in vivo and identify a highly inflammatory, senescent CD24+ osteolineage population cleared by senolytics.

Multiparametric senescent cell phenotyping reveals targets of senolytic therapy in the aged murine skeleton.

Senescence drives organismal aging, yet the deep characterization of senescent cells in vivo remains incomplete. Here, we apply mass cytometry by time-of-flight using carefully validated antibodies to analyze senescent cells at single-cell resolution. We use multiple criteria to identify senescent mesenchymal cells that are growth-arrested and resistant to apoptosis. These p16 + Ki67-BCL-2+ cells are highly enriched for senescence-associated secretory phenotype and DNA damage markers, are strongly associated with age, and their percentages are increased in late osteoblasts/osteocytes and CD24high osteolineage cells. Moreover, both late osteoblasts/osteocytes and CD24high osteolineage cells are robustly cleared by genetic and pharmacologic senolytic therapies in aged mice. Following isolation, CD24+ skeletal cells exhibit growth arrest, senescence-associated ß-galactosidase positivity, and impaired osteogenesis in vitro. These studies thus provide an approach using multiplexed protein profiling to define senescent mesenchymal cells in vivo and identify specific skeletal cell populations cleared by senolytics.

Local senolysis in aged mice only partially replicates the benefits of systemic senolysis.

Clearance of senescent cells (SnCs) can prevent several age-related pathologies, including bone loss. However, the local versus systemic roles of SnCs in mediating tissue dysfunction remain unclear. Thus, we developed a mouse model (p16-LOX-ATTAC) that allowed for inducible SnC elimination (senolysis) in a cell-specific manner and compared the effects of local versus systemic senolysis during aging using bone as a prototype tissue. Specific removal of Sn osteocytes prevented age-related bone loss at the spine, but not the femur, by improving bone formation without affecting osteoclasts or marrow adipocytes. By contrast, systemic senolysis prevented bone loss at the spine and femur and not only improved bone formation, but also reduced osteoclast and marrow adipocyte numbers. Transplantation of SnCs into the peritoneal cavity of young mice caused bone loss and also induced senescence in distant host osteocytes. Collectively, our findings provide proof-of-concept evidence that local senolysis has health benefits in the context of aging, but, importantly, that local senolysis only partially replicates the benefits of systemic senolysis. Furthermore, we establish that SnCs, through their senescence-associated secretory phenotype (SASP), lead to senescence in distant cells. Therefore, our study indicates that optimizing senolytic drugs may require systemic instead of local SnC targeting to extend healthy aging.

In vitro and in vivo effects of zoledronate on senescence and senescence-associated secretory phenotype markers.

In addition to reducing fracture risk, zoledronate has been found in some studies to decrease mortality in humans and extend lifespan and healthspan in animals. Because senescent cells accumulate with aging and contribute to multiple co-morbidities, the non-skeletal actions of zoledronate could be due to senolytic (killing of senescent cells) or senomorphic (inhibition of the secretion of the senescence-associated secretory phenotype [SASP]) actions. To test this, we first performed in vitro senescence assays using human lung fibroblasts and DNA repair-deficient mouse embryonic fibroblasts, which demonstrated that zoledronate killed senescent cells with minimal effects on non-senescent cells. Next, in aged mice treated with zoledronate or vehicle for 8 weeks, zoledronate significantly reduced circulating SASP factors, including CCL7, IL-1ß, TNFRSF1A, and TGFß1 and improved grip strength. Analysis of publicly available RNAseq data from CD115+ (CSF1R/c-fms+) pre-osteoclastic cells isolated from mice treated with zoledronate demonstrated a significant downregulation of senescence/SASP genes (SenMayo). To establish that these cells are potential senolytic/senomorphic targets of zoledronate, we used single cell proteomic analysis (cytometry by time of flight [CyTOF]) and demonstrated that zoledronate significantly reduced the number of pre-osteoclastic (CD115+/CD3e-/Ly6G-/CD45R-) cells and decreased protein levels of p16, p21, and SASP markers in these cells without affecting other immune cell populations. Collectively, our findings demonstrate that zoledronate has senolytic effects in vitro and modulates senescence/SASP biomarkers in vivo . These data point to the need for additional studies testing zoledronate and/or other bisphosphonate derivatives for senotherapeutic efficacy.

In vitro and in vivo effects of zoledronic acid on senescence and senescence-associated secretory phenotype markers.

In addition to reducing fracture risk, zoledronic acid has been found in some studies to decrease mortality in humans and extend lifespan and healthspan in animals. Because senescent cells accumulate with aging and contribute to multiple co-morbidities, the non-skeletal actions of zoledronic acid could be due to senolytic (killing of senescent cells) or senomorphic (inhibition of the secretion of the senescence-associated secretory phenotype [SASP]) actions. To test this, we first performed in vitro senescence assays using human lung fibroblasts and DNA repair-deficient mouse embryonic fibroblasts, which demonstrated that zoledronic acid killed senescent cells with minimal effects on non-senescent cells. Next, in aged mice treated with zoledronic acid or vehicle for 8 weeks, zoledronic acid significantly reduced circulating SASP factors, including CCL7, IL-1ß, TNFRSF1A, and TGFß1 and improved grip strength. Analysis of publicly available RNAseq data from CD115+ (CSF1R/c-fms+) pre-osteoclastic cells isolated from mice treated with zoledronic acid demonstrated a significant downregulation of senescence/SASP genes (SenMayo). To establish that these cells are potential senolytic/senomorphic targets of zoledronic acid, we used single cell proteomic analysis (cytometry by time of flight [CyTOF]) and demonstrated that zoledronic acid significantly reduced the number of pre-osteoclastic (CD115+/CD3e-/Ly6G-/CD45R-) cells and decreased protein levels of p16, p21, and SASP markers in these cells without affecting other immune cell populations. Collectively, our findings demonstrate that zoledronic acid has senolytic effects in vitro and modulates senescence/SASP biomarkers in vivo. These data point to the need for additional studies testing zoledronic acid and/or other bisphosphonate derivatives for senotherapeutic efficacy.