Background In this study, we aimed to evaluate the effects of tocilizumab in adult patients admitted to hospital with COVID-19 with both hypoxia and systemic inflammation. Methods This randomised, controlled, open-label, platform trial (Randomised Evaluation of COVID-19 Therapy [RECOVERY]), is assessing several possible treatments in patients hospitalised with COVID-19 in the UK. Those trial participants with hypoxia (oxygen saturation <92% on air or requiring oxygen therapy) and evidence of systemic inflammation (C-reactive protein ≥75 mg/L) were eligible for random assignment in a 1:1 ratio to usual standard of care alone versus usual standard of care plus tocilizumab at a dose of 400 mg–800 mg (depending on weight) given intravenously. A second dose could be given 12–24 h later if the patient's condition had not improved. The primary outcome was 28-day mortality, assessed in the intention-to-treat population. The trial is registered with ISRCTN (50189673) and ClinicalTrials.gov ( NCT04381936 ). Findings Between April 23, 2020, and Jan 24, 2021, 4116 adults of 21 550 patients enrolled into the RECOVERY trial were included in the assessment of tocilizumab, including 3385 (82%) patients receiving systemic corticosteroids. Overall, 621 (31%) of the 2022 patients allocated tocilizumab and 729 (35%) of the 2094 patients allocated to usual care died within 28 days (rate ratio 0·85; 95% CI 0·76–0·94; p=0·0028). Consistent results were seen in all prespecified subgroups of patients, including those receiving systemic corticosteroids. Patients allocated to tocilizumab were more likely to be discharged from hospital within 28 days (57% vs 50%; rate ratio 1·22; 1·12–1·33; p<0·0001). Among those not receiving invasive mechanical ventilation at baseline, patients allocated tocilizumab were less likely to reach the composite endpoint of invasive mechanical ventilation or death (35% vs 42%; risk ratio 0·84; 95% CI 0·77–0·92; p<0·0001). Interpretation In hospitalised COVID-19 patients with hypoxia and systemic inflammation, tocilizumab improved survival and other clinical outcomes. These benefits were seen regardless of the amount of respiratory support and were additional to the benefits of systemic corticosteroids. Funding UK Research and Innovation (Medical Research Council) and National Institute of Health Research.

SUMMARY Despite a wealth of clinical data showing an association between inflammation and degenerative disorders in elderly, the immune sensors that causally link systemic inflammation to aging remain unclear. Here we detail a mechanism that the Nlrp3 inflammasome controls systemic low grade age-related ‘sterile’ inflammation in both periphery and brain independently of the non-canonical caspase-11 inflammasome. Ablation of Nlrp3 inflammasome protected mice from age-related increases in the innate immune activation, alterations in CNS transcriptome and astrogliosis. Consistent with the hypothesis that systemic low grade inflammation promotes age-related degenerative changes, the deficient Nlrp3 inflammasome mediated caspase-1 activity improved glycemic control and attenuated bone loss and thymic demise. Notably, IL-1 mediated only Nlrp3 inflammasome dependent improvement in cognitive function and motor performance in aged mice. These studies reveal Nlrp3 inflammasome as an upstream target that controls age-related inflammation and offer innovative therapeutic strategy to lower Nlrp3 activity to delay multiple age-related chronic diseases.

Estrogen deficiency is a major risk factor for osteoporosis that is associated with bone inflammation and resorption. Half of women over the age of 50 will experience an osteoporosis related fracture in their lifetime, thus novel therapies are needed to combat post-menopausal bone loss. Recent studies suggest an important role for gut-bone signaling pathways and the microbiota in regulating bone health. Given that the bacterium Lactobacillus reuteri ATCC PTA 6475 (L. reuteri) secretes beneficial immunomodulatory factors, we examined if this candidate probiotic could reduce bone loss associated with estrogen deficiency in an ovariectomized (Ovx) mouse menopausal model. Strikingly, L. reuteri treatment significantly protected Ovx mice from bone loss. Osteoclast bone resorption markers and activators (Trap5 and RANKL) as well as osteoclastogenesis are significantly decreased in L. reuteri treated mice. Consistent with this, L. reuteri suppressed Ovx-induced increases in bone marrow CD4+ T-lymphocytes (which promote osteoclastogenesis) and directly suppressed osteoclastogenesis in vitro. We also identif ied that L. reuteri treatment modifies microbial communities in the Ovx mouse gut. Together, our studies demonstrate that L. reuteri treatment suppresses bone resorption and loss associated with estrogen deficiency. Thus, L. reuteri treatment may be a straightforward and cost-effective approach to reduce post-menopausal bone loss.

The AML/CBFA family of runt homology domain (rhd) transcription factors regulates expression of mammalian genes of the hematopoietic lineage. AML1, AML2 and AML3 are the three AML genes identified to date which influence myeloid cell growth and differentiation. Recently AML-related proteins were identified in an osteoblast-specific promoter binding complex that functionally modulates bone-restricted transcription of the osteocalcin gene. In the present study we demonstrate that in primary rat osteoblasts AML-3 is the AML family member present in the osteoblast-specific complex. Antibody specific for AML-3 completely supershifts this complex, in contrast to antibodies with specificity for AML-1 or AML-2, AML-3 is present as a single 5.4 kb transcript in bone tissues. To establish the functional involvement of AML factors in osteoblast differentiation, we pursued antisense strategies to alter expression of rhd genes. Treatment of osteoblast cultures with rhd antisense oligonucleotides significantly decreased three parameters which are linked to differentiation of normal diploid osteoblasts: the representation of alkaline phosphatase-positive cells, osteocalcin production, and the formation of mineralized nodules. Our findings indicate that AML-3 is a key transcription factor in bone cells and that the activity of rhd proteins is required for completion of osteoblast differentiation.

The first clinical trial of an implantable microchip-based drug delivery device is discussed. Human parathyroid hormone fragment (1-34) ] was delivered from the device in vivo. hPTH(1-34) is the only approved anabolic osteoporosis treatment, but requires daily injections, making patient compliance an obstacle to effective treatment. Furthermore, a net increase in bone mineral density requires intermittent or pulsatile hPTH(1-34) delivery, a challenge for implantable drug delivery products. The microchip-based devices, containing discrete doses of lyophilized hPTH(1-34), were implanted in eight osteoporotic postmenopausal women for 4 months and wirelessly programmed to release doses from the device once daily for up to 20 days. A computer-based programmer, operating in the Medical Implant Communications Service band, established a bidirectional wireless communication link with the implant to program the dosing schedule and receive implant status confirming proper operation. Each woman subsequently received hPTH(1-34) injections in escalating doses. The pharmacokinetics, safety, tolerability, and bioequivalence of hPTH(1-34) were assessed. Device dosing produced similar pharmacokinetics to multiple injections and had lower coefficients of variation. Bone marker evaluation indicated that daily release from the device increased bone formation. There were no toxic or adverse events due to the device or drug, and patients stated that the implant did not affect quality of life.

Decreased bone mass, osteoporosis, and increased fracture rates are common skeletal complications in patients with insulin-dependent diabetes mellitus (IDDM; type I diabetes). IDDM develops from little or no insulin production and is marked by elevated blood glucose levels and weight loss. In this study we use a streptozotocin-induced diabetic mouse model to examine the effect of type I diabetes on bone. Histology and microcomputed tomography demonstrate that adult diabetic mice, exhibiting increased plasma glucose and osmolality, have decreased trabecular bone mineral content compared with controls. Bone resorption could not completely account for this effect, because resorption markers (tartrate-resistant acid phosphatase 5b, urinary deoxypyridinoline excretion, and tartrate-resistant acid phosphatase 5 mRNA) are unchanged or reduced at 2 and/or 4 wk after diabetes induction. However, osteocalcin mRNA (a marker of late-stage osteoblast differentiation) and dynamic parameters of bone formation were decreased in diabetic tibias, whereas osteoblast number and runx2 and alkaline phosphatase mRNA levels did not differ. These findings suggest that the final stages of osteoblast maturation and function are suppressed. We also propose a second mechanism contributing to diabetic bone loss: increased marrow adiposity. This is supported by increased expression of adipocyte markers [peroxisome proliferator-activated receptor gamma2, resistin, and adipocyte fatty acid binding protein (alphaP2)] and the appearance of lipid-dense adipocytes in diabetic tibias. In contrast to bone marrow, adipose stores at other sites are depleted in diabetic mice, as indicated by decreased body, liver, and peripheral adipose tissue weights. These findings suggest that IDDM contributes to bone loss through changes in marrow composition resulting in decreased mature osteoblasts and increased adipose accumulation.

Insulin-dependent diabetes mellitus (IDDM) is associated with increased risk of osteopenia/osteoporosis in humans. The mechanisms accounting for diabetic bone loss remain unclear. Pharmacologic inducers of IDDM, such as streptozotocin, mimic key aspects of diabetes in rodents, allow analysis at the onset of diabetes, and induce diabetes in genetically modified mice. However, side effects of streptozotocin, unrelated to diabetes, can complicate data interpretation. The nonobese diabetic (NOD) mouse model develops diabetes spontaneously without external influences, negating side effects of inducing agents. Unfortunately, in this model the onset of diabetes is unpredictable, occurs in a minority of male mice, and can only be studied in a single mouse strain. To validate the relevance of the more flexible streptozotocin-induced diabetes model for studying diabetes-associated bone loss, we compared its phenotype to the spontaneously diabetic NOD model. Both models exhibited hyperglycemia and loss of body, fat pad, and muscle weight. Furthermore, these genetically different and distinct models of diabetes induction demonstrated similar bone phenotypes marked by significant trabecular bone loss and increased bone marrow adiposity. Correspondingly, both diabetic models exhibited decreased osteocalcin mRNA and increased adipocyte fatty acid-binding protein 2 mRNA levels in isolated tibias and calvaria. Taken together, multiple streptozotocin injection-induced diabetes is a valid model for understanding the acute and chronic pathophysiologic responses to diabetes and their mechanisms in bone.

Osteoporosis can result from intestinal inflammation, as is seen with inflammatory bowel disease. Probiotics, microorganisms that provide a health benefit to the host when ingested in adequate amounts, can have anti-inflammatory properties and are currently being examined to treat inflammatory bowel disease. Here, we examined if treating healthy male mice with Lactobacillus reuteri ATCC PTA 6475 (a candidate probiotic with anti-TNFα activity) could affect intestinal TNFα levels and enhance bone density. Adult male mice were given L. reuteri 6475 orally by gavage for 3 ×/week for 4 weeks. Examination of jejunal and ileal RNA profiles indicates that L. reuteri suppressed basal TNFα mRNA levels in the jejunum and ileum in male mice, but surprisingly not in female mice. Next, we examined bone responses. Micro-computed tomography demonstrated that L. reuteri 6475 treatment increased male trabecular bone parameters (mineral density, bone volume fraction, trabecular number, and trabecular thickness) in the distal femur metaphyseal region as well as in the lumbar vertebrae. Cortical bone parameters were unaffected. Dynamic and static histomorphometry and serum remodeling parameters indicate that L. reuteri ingestion increases osteoblast serum markers and dynamic measures of bone formation in male mice. In contrast to male mice, L. reuteri had no effect on bone parameters in female mice. Taken together our studies indicate that femoral and vertebral bone formation increases in response to oral probiotic use, leading to increased trabecular bone volume in male mice.