miR-29b inhibits TGF-β1-induced cell proliferation in articular chondrocytes

Transforming growth factor β1 (TGF-β1) is a known regulator of chondrocyte proliferation and promotes cartilage repair in osteoarthritis (OA). microRNA-29b-3p (miR-29b-3p) is downregulated by TGF-β1 and overexpressed in OA cartilage. However, the ability of miR-29b-3p to mediate the chondrocyte pro-proliferative effects of TGF-β1 is not yet understood. This current study aimed to investigate the effect of miR-29b-3p on TGF-β1-induced cell proliferation in murine articular chondrocytes. The stimulation of chondrocytes by TGF-β1 for 24 h resulted in the downregulation of miR-29b-3p expression. The ratio of G0/G1 phase cells decreased in response to TGF-β1 whereas the ratio of S phase cells was increased. Consistent with this observation, miR-29b-3p overexpression inhibited TGF-β1’s ability to promote the ratio of S phase cells and downregulate the ratio of G0/G1 phase cells. These findings suggest that the downregulation of miR-29b-3p is a likely requirement for TGF-β1-mediated proliferation of murine articular chondrocytes. Furthermore, implying that miR-29b-3p expression may be involved in reduced chondrocyte proliferation in OA

Ablation of Enpp6 results in transient bone hypomineralization

C.F. was supported by the Biotechnology and Biological Sciences Research Council (BBSRC) via an Institute Strategic Programme Grant Funding (BB/J004316/1). S.D. was supported through a BBSRC EASTBIO Doctoral Training Partnership studentship award (1803936) and N.M.M. was supported by a Wellcome Trust New Investigator Award (100981/Z/13/Z). S.D. wrote the manuscript. S.D., K.S., S-N.H., and L.A.S. carried out experimental work. W.P.C., R.W. and N.M.M. provided reagents and materials. A.J.S., F.N. and C.F. contributed to conceptualization of the study and experimental design. All authors reviewed and edited the manuscript and approved the final version. All authors state that they have no conflicts of interest.Biomineralization is a fundamental process key to the development of the skeleton. The phosphatase orphan phosphatase 1 (PHOSPHO1), which likely functions within extracellular matrix vesicles, has emerged as a critical regulator of biomineralization. The biochemical pathways which generate intravesicular PHOSPHO1 substrates are however currently unknown. We hypothesized that the enzyme ectonucleotide pyrophosphatase/phosphodiesterase (ENPP6) is an upstream source of PHOSPHO1 substrate. To test this, we characterized skeletal phenotypes of mice homozygous for a targeted deletion of Enpp6 (Enpp6‒/‒). Micro-computed tomography of the trabecular compartment revealed transient hypomineralization in Enpp6‒/‒ tibiae (p 0.01) and osteoid surface (p < 0.05) which recovered by 12 weeks but was not accompanied by changes in osteoblast or osteoclast number. This study is the first to characterize the skeletal phenotype of Enpp6‒/‒ mice, revealing transient hypomineralization in young animals compared to wild-type controls. These data suggest that ENPP6 is important for bone mineralization and may function upstream of PHOSPHO1 as a novel means of generating its substrates inside matrix vesicles.Publisher PDFPeer reviewe

Mitochondrial dysfunction and mitophagy blockade contribute to renal osteodystrophy in chronic kidney disease-mineral bone disorder

Chronic kidney disease–mineral and bone disorder (CKD-MBD) presents with extra-skeletal calcification and renal osteodystrophy (ROD). The origins of ROD likely lie with elevated uremic toxins and/or an altered hormonal profile but the cellular events responsible remain unclear. Here, we report that stalled mitophagy contributes to mitochondrial dysfunction in bones of a CKD-MBD mouse model, and also human CKD-MBD patients. RNA-seq analysis exposed an altered expression of genes associated with mitophagy and mitochondrial function in tibia of CKD-MBD mice. The accumulation of damaged osteocyte mitochondria and the expression of mitophagy regulators, p62/SQSTM1, ATG7 and LC3 was inconsistent with functional mitophagy, and in mito-QC reporter mice with CKD-MBD, there was a 2.3-fold increase in osteocyte mitolysosomes. Altered expression of mitophagy regulators in human CKD-MBD bones was also observed. To determine if uremic toxins were possibly responsible for these observations, indoxyl sulfate treatment of osteoblasts revealed mitochondria with distorted morphology and whose membrane potential and oxidative phosphorylation were decreased, and oxygen-free radical production increased. The altered p62/SQSTM1 and LC3-II expression was consistent with impaired mitophagy machinery and the effects of indoxyl sulfate were reversible by rapamycin. In conclusion, mitolysosome accumulation from impaired clearance of damaged mitochondria may contribute to the skeletal complications, characteristic of ROD. Targeting mitochondria and the mitophagy process may therefore offer novel routes for intervention to preserve bone health in patients with ROD. Such approaches would be timely as our current armamentarium of anti-fracture medications has not been developed for, or adequately studied in, patients with severe CKD-MBD

Acute kidney disease following COVID-19 vaccination: a single-center retrospective study

BackgroundRare cases of de novo or relapsed kidney diseases associated with vaccination against coronavirus disease 2019 (COVID-19) have been increasingly reported. The aim of this study was to report the incidence, etiologies, and outcomes of acute kidney disease (AKD) following COVID-19 vaccination.MethodsThis retrospective study extracted cases from renal registry of a single medical center from 1 March 2021 to 30 April 2022, prior to the significant surge in cases of the Omicron variant of COVID-19 infection in Taiwan. Adult patients who developed AKD after COVID-19 vaccination were included. We utilized the Naranjo score as a causality assessment tool for adverse vaccination reactions and charts review by peer nephrologists to exclude other causes. The etiologies, characteristics, and outcomes of AKD were examined.ResultsTwenty-seven patients (aged 23 to 80 years) with AKD were identified from 1,897 vaccines (estimated rate of 13.6 per 1000 patient-years within the renal registry). A majority (77.8%) of vaccine received messenger RNA-based regimens. Their median (IQR) Naranjo score was 8 (6-9) points, while 14 of them (51.9%) had a definite probability (Naranjo score ≥ 9). The etiologies of AKD included glomerular disease (n = 16) consisting of seven IgA nephropathy, four anti-neutrophil cytoplasmic antibodies-associated glomerulonephritis (AAN), three membranous glomerulonephritis, two minimal change diseases, and chronic kidney disease (CKD) with acute deterioration (n = 11). Extra-renal manifestations were found in four patients. Over a median (IQR) follow-up period of 42 (36.5–49.5) weeks, six patients progressed to end-stage kidney disease (ESKD).ConclusionBesides glomerulonephritis (GN), the occurrence of AKD following COVID-19 vaccination may be more concerning in high-risk CKD patients receiving multiple doses. Patients with the development of de novo AAN, concurrent extra-renal manifestations, or pre-existing moderate to severe CKD may exhibit poorer kidney prognosis

Mechanisms underpinning chronic kidney disease-mineral and bone disorder

Chronic kidney disease (CKD) is an irreversible systemic disease characterised by the gradual loss of kidney function over time. Patients with progressive CKD frequently encounter altered levels of circulating and bone-derived hormones which can dysregulate mineral metabolism leading to ectopic calcification and compromised bone formation. Generally, CKD presents with increased levels of parathyroid hormone (PTH), fibroblastic growth factor-23 (FGF23), and phosphate (Pi), all of which are able to affect bone homeostasis through direct and indirect effects on bone-forming and resorbing cells. FGF23 is a phosphaturic factor that starts to increase in the early stages of CKD. It controls Pi homeostasis and impacts bone mineralisation by inhibiting tissue nonspecific alkaline phosphatase (TNAP) and stimulating pyrophosphate (PPi) expression in bone. PTH is a key regulator of bone remodelling and has biphasic effects on bone metabolism. While intermittent PTH stimulation facilitates bone formation, chronic exposure to PTH as observed in advanced CKD results in bone loss. Besides, high Pi levels that emerge at the latter stages of CKD may aggravate skeletal mineralisation by increasing bone resorption through the promotion of FGF23 and PTH secretion and their associated transcriptional effectors. These observations led to the hypotheses that FGF23, PTH and Pi may possess a “direct effect” on bone cells that mediate bone remodelling and mineralisation. Although growing evidence indicates that FGF23, PTH and Pi modulate bone matrix mineralisation, it is also possible however that the altered endocrine milieu directly targets the expression of key phosphatases that are critical for skeletal mineralisation. Two of the most widely studied phosphatases involved in skeletal mineralisation are PHOSPHO1 and TNAP. During mineralisation, PHOSPHO1 liberates Pi to be incorporated into the mineral phase through hydrolysis of its membrane-bound substrates leading to an increase in the Pi/PPi ratio within matrix vesicles (MVs). TNAP, highly expressed on the membranes of MVs hydrolyses PPi to facilitate the propagation of hydroxyapatite (HA) in the extracellular matrix (ECM). A complete absence of ECM mineralisation is observed in PHOSHO1; TNAP double knock-out (Phospho1-/-; Alpl-/-) mice. Despite clear links between TNAP and PHOSPHO1 in the control of skeletal mineralisation, there is a lack of evidence about the expression profiles of PHOSPHO1 and TNAP in mineral and bone disorders (MBD) in CKD. The work described in this thesis characterised the expression of PHOSPHO1, TNAP, and other key genes associated with ECM mineralisation in in vitro models of CKD by continuously exposing murine osteoblasts to FGF23, PTH and Pi and in vivo murine model of CKD-MBD. In the adenine-induced murine model of CKD, mice presented with modest skeletal complications that were characteristic of renal osteodystrophy (ROD), a feature of CKD-MBD. Notably, BMD of trabecular bone was decreased whereas it was increased in cortical bone of CKD-MBD mice. These changes in CKD bones were accompanied by decreased TNAP and PHOSPHO1 expression. However, cortical bone BMD was unchanged in Phospho1 knockout (P1KO) CKD mice suggesting that the increased cortical BMD noted in CKD was driven by the increased PHOSPHO1 expression. The administration of Pi, PTH, and FGF23 in vitro had various effects on osteoblast ECM mineralisation but all three decreased PHOSPHO1 and TNAP expression in culture. Overall, the in vivo and in vitro experimental models of CKD were the first to implicate PHOSPHO1 function in the altered mineralisation status of CKD bones. Furthermore, this thesis provides the first detailed transcriptomic analyses (RNA-seq) of bone from a murine CKD-MBD model which will prove invaluable in understanding the altered bone metabolism and mineralisation noted during disease progression. Intriguingly, mitochondrial dysfunction was identified as a possible causative pathological mechanism implicated in altered bone remodelling of CKD-MBD. This is a novel observation. Indoxyl sulfate (IS), is a representative renal toxin and acts as a bone toxin by inducing reactive oxygen species (ROS) overproduction inside the cell. This work revealed an extensive characterisation of the effects of IS on osteoblast mitochondrial mass, ROS production, and energy metabolism in primary osteoblasts. The pharmacological mitophagy activator, rapamycin can restore the PARKIN-associated mitophagy pathways mediating the effects of IS on mitochondria. Finally, in vivo models such as mito-QC CKD reporter mice provide direct evidence of mitophagy abnormalities in the bone of CKD-MBD mice. In addition, the reduction of ATG7 (a protein essential for autophagy) in CKD human femurs supports the hypothesis that defective autophagy in CKD bone contributes to mitophagy dysfunction. Such findings will help to identify promising therapeutic options to manage the skeletal complications of CKD encountered in clinical practice

A fatal case of drug reaction with eosinophilia and systemic symptom syndrome associated with cytomegalovirus reactivation

Drug reaction with eosinophilia and systemic symptom (DRESS) syndrome is a severe adverse drug-induced reaction. Diagnosing DRESS syndrome is also challenging due to the diversity of cutaneous eruption and the organs involved. Here, we described an 88-year-old Chinese woman who developed DRESS syndrome under combined therapy of nonsteroidal anti-inflammatory drugs (NSAIDs) and chlormezanone (CM) accompanied with cytomegalovirus reactivation. DRESS syndrome should be highly suspected in patients with symptoms, including skin rash, fever, liver involvement, hypereosinophilia, and lymphadenopathy, especially coexisted with reactivation of the cytomegalovirus. Early withdrawal of the culprit drug is necessary once the diagnosis is established