microRNA-181a-5p antisense oligonucleotides attenuate osteoarthritis in facet and knee joints

ObjectivesWe recently identified microRNA-181a-5p (miR-181a-5p) as a critical mediator involved in the destruction of lumbar facet joint (FJ) cartilage. In this study, we tested if locked nucleic acid (LNA) miR-181a-5p antisense oligonucleotides (ASO) could be used as a therapeutic to limit articular cartilage degeneration.MethodsWe used a variety of experimental models consisting of both human samples and animal models of FJ and knee osteoarthritis (OA) to test the effects of LNA-miR-181a-5p ASO on articular cartilage degeneration. Histopathological analysis including immunohistochemistry and in situ hybridisation were used to detect key OA catabolic markers and microRNA, respectively. Apoptotic/cell death markers were evaluated by flow cytometry. qPCR and immunoblotting were applied to quantify gene and protein expression.ResultsmiR-181a-5p expression was increased in human FJ OA and knee OA cartilage as well as injury-induced FJ OA (rat) and trauma-induced knee OA (mouse) cartilage compared with control cartilage, correlating with classical OA catabolic markers in human, rat and mouse cartilage. We demonstrated that LNA-miR-181a-5p ASO in rat and mouse chondrocytes reduced the expression of cartilage catabolic and chondrocyte apoptotic/cell death markers in vitro. Treatment of OA-induced rat FJ or mouse knee joints with intra-articular injections of in vivo grade LNA-miR-181a-5p ASO attenuated cartilage destruction, and the expression of catabolic, hypertrophic, apoptotic/cell death and type II collagen breakdown markers. Finally, treatment of LNA-miR-181a-5p ASO in cultures of human knee OA chondrocytes (in vitro) and cartilage explants (ex vivo) further demonstrated its cartilage protective effects.ConclusionsOur data demonstrate, for the first time, that LNA-miR-181a-5p ASO exhibit cartilage-protective effects in FJ and knee OA.

Download Full-text

KUS121 attenuates the progression of monosodium iodoacetate-induced osteoarthritis in rats

Scientific Reports ◽

10.1038/s41598-021-95173-6 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Sachiko Iwai ◽

Hanako O. Ikeda ◽

Hisashi Mera ◽

Kohei Nishitani ◽

Motoo Saito ◽

...

Keyword(s):

Cell Death ◽

Er Stress ◽

Apoptotic Cell ◽

Intraperitoneal Administration ◽

Atp Depletion ◽

Knee Joints ◽

Cellular Protection ◽

Monosodium Iodoacetate

AbstractCurrently there is no effective treatment available for osteoarthritis (OA). We have recently developed Kyoto University Substances (KUSs), ATPase inhibitors specific for valosin-containing protein (VCP), as a novel class of medicine for cellular protection. KUSs suppressed intracellular ATP depletion, endoplasmic reticulum (ER) stress, and cell death. In this study, we investigated the effects of KUS121 on chondrocyte cell death. In cultured chondrocytes differentiated from ATDC5 cells, KUS121 suppressed the decline in ATP levels and apoptotic cell death under stress conditions induced by TNFα. KUS121 ameliorated TNFα-induced reduction of gene expression in chondrocytes, such as Sox9 and Col2α. KUS121 also suppressed ER stress and cell death in chondrocytes under tunicamycin load. Furthermore, intraperitoneal administration of KUS121 in vivo suppressed chondrocyte loss and proteoglycan reduction in knee joints of a monosodium iodoacetate-induced OA rat model. Moreover, intra-articular administration of KUS121 more prominently reduced the apoptosis of the affected chondrocytes. These results demonstrate that KUS121 protects chondrocytes from stress-induced cell death in vitro and in vivo, and indicate that KUS121 is a promising novel therapeutic agent to prevent the progression of OA.

Download Full-text

The effect of continuous combined 17β-oestradiol and dihydrodydrogesterone on apoptotic cell death and proliferation of human breast cancer cells in vitro

European Journal of Cancer ◽

10.1016/s0959-8049(02)00293-9 ◽

2002 ◽

Vol 38 ◽

pp. 69-70 ◽

Cited By ~ 6

Author(s):

H.R Franke ◽

I Vermes

Keyword(s):

Breast Cancer ◽

Cell Death ◽

Cancer Cells ◽

Human Breast Cancer ◽

Breast Cancer Cells ◽

Apoptotic Cell ◽

Apoptotic Cell Death ◽

Human Breast Cancer Cells ◽

Human Breast

Download Full-text

Apoptotic signaling through CD95 (Fas/Apo-1) activates an acidic sphingomyelinase.

Journal of Experimental Medicine ◽

10.1084/jem.180.4.1547 ◽

1994 ◽

Vol 180 (4) ◽

pp. 1547-1552 ◽

Cited By ~ 373

Author(s):

M G Cifone ◽

R De Maria ◽

P Roncaioli ◽

M R Rippo ◽

M Azuma ◽

...

Keyword(s):

Cell Death ◽

Cell Lines ◽

Intracellular Signaling ◽

Apoptotic Cell ◽

Human Tumor ◽

Apoptotic Cell Death ◽

Cell Surface Receptor ◽

Cross Linking ◽

Cell Extracts

Intracellular pathways leading from membrane receptor engagement to apoptotic cell death are still poorly characterized. We investigated the intracellular signaling generated after cross-linking of CD95 (Fas/Apo-1 antigen), a broadly expressed cell surface receptor whose engagement results in triggering of cellular apoptotic programs. DX2, a new functional anti-CD95 monoclonal antibody was produced by immunizing mice with human CD95-transfected L cells. Crosslinking of CD95 with DX2 resulted in the activation of a sphingomyelinase (SMase) in promyelocytic U937 cells, as well as in other human tumor cell lines and in CD95-transfected murine cells, as demonstrated by induction of in vivo sphingomyelin (SM) hydrolysis and generation of ceramide. Direct in vitro measurement of enzymatic activity within CD95-stimulated U937 cell extracts, using labeled SM vesicles as substrates, showed strong SMase activity, which required pH 5.0 for optimal substrate hydrolysis. Finally, all CD95-sensitive cell lines tested could be induced to undergo apoptosis after exposure to cell-permeant C2-ceramide. These data indicate that CD95 cross-linking induces SM breakdown and ceramide production through an acidic SMase, thus providing the first information regarding early signal generation from CD95, and may be relevant in defining the biochemical nature of intracellular messengers leading to apoptotic cell death.

Download Full-text

Dapagliflozin Alleviates Renal Fibrosis by Inhibiting RIP1-RIP3-MLKL-Mediated Necroinflammation in Unilateral Ureteral Obstruction

Frontiers in Pharmacology ◽

10.3389/fphar.2021.798381 ◽

2022 ◽

Vol 12 ◽

Author(s):

Mei Ying Xuan ◽

Shang Guo Piao ◽

Jun Ding ◽

Qi Yan Nan ◽

Mei Hua Piao ◽

...

Keyword(s):

Cell Death ◽

Ureteral Obstruction ◽

Renal Fibrosis ◽

Apoptotic Cell ◽

Subsequent Development ◽

Unilateral Ureteral Obstruction ◽

Apoptotic Cell Death ◽

Inflammasome Activation ◽

Renal Tubular

Dapagliflozin, a sodium-glucose cotransporter-2 inhibitor, offers renoprotection in diabetes. However, potential for use in nondiabetic kidney disease remains unknown. Herein, we assessed whether dapagliflozin alleviates renal fibrosis by interfering with necroinflammation in a rat model of unilateral ureteral obstruction (UUO) and in vitro. After induction of UUO, rats were administered dapagliflozin daily for seven consecutive days. UUO induced significant renal tubular necrosis and overexpression of RIP1-RIP3-MLKL axis proteins; these coincided with NLRP3 inflammasome activation, and subsequent development of renal fibrosis. Oxidative stress caused by UUO is tightly associated with endoplasmic reticulum stress and mitochondrial dysfunction, leading to apoptotic cell death through Wnt3α/β-catenin/GSK-3β signaling; all of which were abolished by both dapagliflozin and specific RIP inhibitors (necrostatin-1 and GSK872). In H2O2-treated HK-2 cells, dapagliflozin and RIP inhibitors suppressed overexpression of RIP1-RIP3-MLKL proteins and pyroptosis-related cytokines, decreased intracellular reactive oxygen species production and apoptotic cell death, whereas cell viability was improved. Moreover, activated Wnt3α/β-catenin/GSK-3β signaling was inhibited by dapagliflozin and Wnt/β-catenin inhibitor ICG-001. Our findings suggest that dapagliflozin ameliorates renal fibrosis by inhibiting RIP1-RIP3-MLKL-mediated necroinflammation via Wnt3α/β-catenin/GSK-3β signaling in UUO.

Download Full-text

Inhibition of FAS/FAS ligand-mediated apoptotic cell death of lymphocytes in vitro by circulating anti-FAS ligand autoantibodies in patients with systemic lupus erythematosus

Arthritis & Rheumatism ◽

10.1002/1529-0131(199802)41:2<344::aid-art19>3.0.co;2-j ◽

1998 ◽

Vol 41 (2) ◽

pp. 344-353 ◽

Cited By ~ 36

Author(s):

Noboru Suzuki ◽

Motohide Ichino ◽

Shoji Mihara ◽

Sakae Kaneko ◽

Tsuyoshi Sakane

Keyword(s):

Systemic Lupus Erythematosus ◽

Cell Death ◽

Lupus Erythematosus ◽

Fas Ligand ◽

Apoptotic Cell ◽

Apoptotic Cell Death ◽

Systemic Lupus

Download Full-text

Biochemical Characterisation of an In Vitro Model of Hepatocellular Apoptotic Cell Death

Alternatives to Laboratory Animals ◽

10.1177/026119290903700210 ◽

2009 ◽

Vol 37 (2) ◽

pp. 209-218 ◽

Cited By ~ 10

Author(s):

Mathieu Vinken ◽

Elke Decrock ◽

Elke De Vuyst ◽

Luc Leybaert ◽

Tamara Vanhaecke ◽

...

Keyword(s):

Cell Death ◽

In Vitro Model ◽

Caspase 3 ◽

Fas Ligand ◽

Apoptotic Cell ◽

Annexin V ◽

Apoptotic Cell Death ◽

Biochemical Characterisation ◽

Vitro Model

This study was set up to critically evaluate a commonly-used in vitro model of hepatocellular apoptotic cell death, in which freshly isolated hepatocytes, cultured in a monolayer configuration, are exposed to a combination of Fas ligand and cycloheximide for six hours. A set of well-acknowledged cell death markers was addressed: a) cell morphology was studied by light microscopy; b) apoptotic and necrotic cell populations were quantified by in situ staining with Annexin-V, Hoechst 33342 and propidium iodide (PI); c) apoptotic and necrotic activities were monitored by probing caspase 3-like activity and measuring the extracellular leakage of lactate dehydrogenase (LDH), respectively; and d) the expression of apoptosis regulators was investigated by immunoblotting. The initiation of apoptosis was evidenced by the activation of caspase 8 and caspase 9, and increased Annexin-V reactivity. Progression through the apoptotic process was confirmed by the activation of caspase 3 and Bid, the enhanced expression of Bax, and the occurrence of nuclear fragmentation. Late transition to a necrotic appearance was demonstrated by an increased number of PI-positive cells and augmented extracellular release of LDH. Thus, the in vitro model allows the study of the entire course of Fas-mediated hepatocellular apoptotic cell death, which is not possible in vivo. This experimental system can serve a broad range of in vitro pharmaco-toxicological purposes, thereby directly assisting in the reduction of animal experimentation.

Download Full-text