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Renal fibrosis, a common pathological manifestation of virtually all types of chronic kidney disease (CKD), ultimately predisposes patients to end-stage renal disease. However, there is no effective therapy for renal fibrosis. Our earlier studies proved that RIP3-mediated necroptosis might be an important mode of renal tubular cell death in rats with chronic renal injury. Under transmission electron microscopy (TEM), we found morphological changes in the necrosis of human renal tissue, and the percentage of necrotic cells increased significantly in patients with stages 2 and 3a CKD. Immunofluorescence analyses showed that the percentages of TUNEL+ /RIP3+ double-positive and TUNEL+ /MLKL+ double-positive tubular epithelial cells in renal tubules of patients with stages 2 and 3a CKD were significantly increased compared to those in control patients without renal disease. Immunohistochemistry analyses of renal biopsy specimens from patients with CKD revealed RIP3, MLKL, and p-MLKL upregulation in patients with stages 2 and 3a CKD, suggesting that necroptosis of renal tubular epithelial cells in CKD patients occurs, and the peak of necroptosis was in stages 2 and 3a CKD. We showed that profibrotic factor proteins (TGF-ß1, Smad2 and Smad3) and fibroblast activation markers (α-SMA and Vimentin) were specifically upregulated in stage 2 and 3a CKD patients. In addition, Pearson correlation analysis showed that the percentage of necroptotic renal tubular epithelial cells was positively correlated with TGF-ß1 and collagen-I. We also showed that RIP1/3 or MLKL inhibitors decreased the expression of RIP3, MLKL, TGF-ß1, and Smad3 in HK-2 cells treated with TNF-α. FGF-2, α-SMA, Vimentin and FN were overexpressed in the hRIFs cultured with the supernatant of necroptotic HK-2 cells, whereas necroptosis blockers (Nec-1s, GSK'872 and NSA) and TGF-ß1/Smad3 pathway antagonists (LY364947 and SIS3) reduced FGF-2, α-SMA, Vimentin and FN levels. Collectively, necroptosis of renal tubular epithelial cells in CKD patients occurs, and the peak of necroptosis was in stages 2 and 3a CKD. Renal tubular epithelial cell necroptosis mediates renal tubulointerstitial fibrosis in patients with chronic kidney disease, which is related to the TGF-ß1/Smad3 signaling pathway.
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Insuficiencia Renal Crónica , Factor de Crecimiento Transformador beta1 , Humanos , Ratas , Animales , Factor de Crecimiento Transformador beta1/metabolismo , Necroptosis , Vimentina/metabolismo , Factor 2 de Crecimiento de Fibroblastos/metabolismo , Fibrosis , Células Epiteliales/metabolismo , Insuficiencia Renal Crónica/metabolismo , Riñón/metabolismo , Necrosis/patologíaRESUMEN
Necroptosis, an alternative mode of programmed cell death, has crucial pathophysiological roles in many diseases, but its effect on chronic kidney disease (CKD) is poorly understood. Therefore, we assessed necroptosis and its pathophysiological effects in a widely used remnant-kidney rat model. We found that necroptotic cell death and the highest level of receptor interaction protein kinase 1 (RIP1) and receptor interaction protein kinase 3 (RIP3), critical signalling molecules for necroptosis, appeared 8 weeks after subtotal nephrectomy (SNX) surgery. After treatment with Necrostatin-1 (Nec-1), renal function and renal pathologic changes were significantly improved; the overexpression of RIP1, RIP3, mixed lineage kinase domain-like (MLKL) and dynamin-related protein 1 (Drp1) was reduced; and necroptosis was inhibited. These results indicated that necroptosis mediated by RIP1 and RIP3 participates in the loss of renal cells of subtotal nephrectomised rats and might be one of main causes of the excessive loss of renal cells during the sustained progression of renal fibrosis.
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Apoptosis , Riñón/enzimología , Riñón/patología , Proteínas Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Insuficiencia Renal Crónica/enzimología , Insuficiencia Renal Crónica/patología , Animales , Células Cultivadas , Imidazoles/uso terapéutico , Indoles/uso terapéutico , Riñón/cirugía , Masculino , Necrosis/metabolismo , Necrosis/patología , Nefrectomía , Ratas , Ratas Sprague-Dawley , Especies Reactivas de Oxígeno/metabolismo , Insuficiencia Renal Crónica/tratamiento farmacológico , Resultado del TratamientoRESUMEN
Postmenopausal osteoporosis (PMOP) is a common kind of osteoporosis that is associated with excessive osteocyte death and bone loss. Previous studies have shown that TNF-α-induced osteocyte necroptosis might exert a stronger effect on PMOP than apoptosis, and TLR4 can also induce cell necroptosis, as confirmed by recent studies. However, little is known about the relationship between TNF-α-induced osteocyte necroptosis and TLR4. In the present study, we showed that TNF-α increased the expression of TLR4, which promoted osteocyte necroptosis in PMOP. In patients with PMOP, TLR4 was highly expressed at skeletal sites where exists osteocyte necroptosis, and high TLR4 expression is correlated with enhanced TNF-α expression. Osteocytes exhibited robust TLR4 expression upon exposure to necroptotic osteocytes in vivo and in vitro. Western blotting and immunofluorescence analyses demonstrated that TNF-α upregulated TLR4 expression in vitro, which might further promote osteocyte necroptosis. Furthermore, inhibition of TLR4 by TAK-242 in vitro effectively blocked osteocyte necroptosis induced by TNF-α. Collectively, these results suggest a novel TLR4-mediated process of osteocyte necroptosis, which might increase osteocyte death and bone loss in the process of PMOP.
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Osteocitos , Osteoporosis Posmenopáusica , Receptor Toll-Like 4 , Factor de Necrosis Tumoral alfa , Femenino , Humanos , Necroptosis , Osteocitos/metabolismo , Osteoporosis Posmenopáusica/metabolismo , Receptor Toll-Like 4/metabolismo , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Background: Wound repair is a new field that has emerged in China in the last 5 years. Exposed tendon wounds are one of the most common problems faced in wound treatment today, as the poor blood supply leads to low survival rates of skin grafts. This paper explores the feasibility of applying the Masquelet technique to repair tendon-exposed wounds. Method: We examined 12 patients with tendon-exposed wounds, 5 males and 7 females, from January 2021 to November 2021, including 2 patients with post-traumatic wounds, 8 diabetic patients with dorsal wounds, and 2 patients with various chronic infections. The Masquelet technique was employed to treat these wounds. The wound surface was sealed with antibiotic bone cement to form an induction membrane, the cement was removed after 3-4 weeks, and the wound was repaired with skin grafts to observe survival, appearance, texture, healing, and related functions. Results: All wounds were covered with antibiotic bone cement, and after 3-4 weeks, an induction membrane was applied, and in 10 out of 12 patients, full-thickness skin grafts were applied, and the patients survived. However, in 2 patients, the skin became partially necrotic, but these patients recovered by changing medications. Conclusion: The current study found that direct skin grafting may effectively treat exposed tendon wounds once the Masquelet approach generates the induction membrane. Further, this method is less difficult, less expensive, and easier to care for the procedure that deserves to be used more frequently.
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Sleep duration suggests some association with osteoporosis and cardiometabolic diseases, but it is unknown if these associations are causal or confounded. In this two-sample Mendelian randomization (MR) study, we included the largest genome-wide association studies (GWASs) associated with sleep duration and the outcome measures of osteoporosis and cardiometabolic diseases. Finally, 25 single nucleotide polymorphisms (SNPs) associated with short sleep duration and 7 SNPs associated with long sleep duration obtained the genome-wide significance (P < 5 × 10-8) and were used as instrumental variables. Genetic predisposition to short sleep duration was strongly associated with increased risk of coronary artery disease (beta-estimate: 0.199, 95% confidence interval CI: 0.081 to 0.317, standard error SE:0.060, P value = 0.001) and heart failure (beta-estimate: 0.145, 95% CI: 0.025 to 0.264, SE:0.061, P value = 0.017), which were both confirmed by the sensitivity analyses. Both short and long sleep duration may reduce the estimated bone mineral density (eBMD, beta-estimate: -0.086, 95% CI: -0.141 to -0.031, SE:0.028, P value = 0.002 for short sleep duration; beta-estimate: -0.080, 95% CI: -0.120 to -0.041, SE:0.020, P value < 0.0001 for long sleep duration). There was limited evidence of associations between sleep duration and fracture, type 2 diabetes, atrial fibrillation, fasting glucose, fasting insulin, or HbA1c. This study provides robust evidence that short sleep duration is causally associated with high risk of coronary artery disease and heart failure and suggests that short sleep duration should be avoided to prevent these two cardiovascular diseases. Short and long sleep duration show some MR association with reduced eBMD, which indicates that both short and long sleep duration may be prevented to reduce the incidence of osteoporosis.
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Enfermedades Cardiovasculares , Enfermedad de la Arteria Coronaria , Diabetes Mellitus Tipo 2 , Insuficiencia Cardíaca , Insulinas , Osteoporosis , Trastornos del Sueño-Vigilia , Humanos , Estudio de Asociación del Genoma Completo , Análisis de la Aleatorización Mendeliana , Diabetes Mellitus Tipo 2/genética , Enfermedad de la Arteria Coronaria/genética , Hemoglobina Glucada/genética , Polimorfismo de Nucleótido Simple/genética , Enfermedades Cardiovasculares/genética , Osteoporosis/genética , Sueño/genética , GlucosaRESUMEN
OBJECTIVE: To observe the apoptosis of spermatogenic cells and the expressions of Bcl-2 and Bax proteins after burying the testis in the inguinal pocket, and to investigate their relationship. METHODS: We randomly divided 36 healthy male New Zealand white rabbits into an experimental group (n = 18) and a control group (n = 18). Models were established by burying testes in the inguinal pocket in the experimental group, while the controls were left untreated. At the end of the 8th week after surgery, 6 animals were randomly taken from each group for measurement of the testis surface temperature and testicular biopsy. The apoptosis of spermatogenic cells in the testis tissues was detected by TUNEL assay, and the expressions of Bcl-2 and Bax proteins determined by immunohistochemistry and imaging analysis. RESULTS: At 8 weeks after burying the testis in the inguinal pocket, the testicular surface temperature was significantly higher in the experimental group than in the control ([ 38.02 +/- 0.36] degrees C vs [36.15 +/- 0.64 ] degrees C, P < 0.05), and so was the apoptosis index (AI) of spermatogenic cells ([89.69 +/- 3.76] % vs [7.73 +/- 4.95 ] %, P < 0.05). The expression of the Bax protein in the testis was significantly increased, while that of the Bcl-2 protein remarkably decreased in the experimental group as compared with the control group (P < 0.05). The apoptotic cells were mostly primary spermatocytes and round spermatids. CONCLUSION: Elevated local temperature of the testis buried in the inguinal pocket increases the apoptosis of spermatogenic cells, and the spermatogenic cell apoptosis is highly correlated with the decreased expression of Bcl-2 and increased expression of Bax. The changes in the expressions of Bcl-2 and Bax proteins were a main mechanism behind the temperature elevation-induced apoptosis of spermatogenic cells.
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Apoptosis , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Espermátides/metabolismo , Testículo/metabolismo , Proteína X Asociada a bcl-2/metabolismo , Animales , Ingle , Masculino , Conejos , Temperatura , Testículo/patologíaRESUMEN
BACKGROUND: Administering corticosteroid is an effective therapeutic strategy for treating most inflammatory conditions. However, there is a chance for corticosteroid treatment to adversely affect bones, resulting in corticosteroid-induced osteoporosis, which is a highly prevalent type of secondary osteoporosis. Elevated bone resorption and reduced formation of bone are pathogenesis indicators of corticosteroid-induced osteoporosis. Preventative therapy is recommended for patients initiating steroids. This study aims to evaluate the efficiency of calcium and vitamin D in treating adults diagnosed with osteoporosis caused by corticosteroid therapy. METHODS: Electronic databases will be searched systematically to source studies that have evaluated the efficiency of calcium and vitamin D as a treatment method for adult patients with osteoporosis from corticosteroid therapy. The databases include, PubMed, EMBASE, the Cochrane Library, Scopus, and Web of Science. The timeline of the search will be limited from inception to November 2020. This study will utilize the Cochrane risk of bias tool to assess the quality of the studies reviewed. Moreover, appropriate methods will be chosen to analyze the data. The RevMan 5.3 software is utilized to perform statistical analysis. RESULTS: This study will provide additional practical and targeted results of evaluating the efficiency of calcium and vitamin D in treating adults with corticosteroid-induced osteoporosis. CONCLUSION: The results of this study will provide further evidence about calcium and vitamin D in treating adults with corticosteroid-induced osteoporosis, clinicians and policymakers can make practical use of the results. ETHICS AND DISSEMINATION: Since this systematic review does not involve any human or animal participants, an ethics approval is not required. SYSTEMATIC REVIEW REGISTRATION: Aug 19, 2021. osf.io/zvb38. (https://osf.io/zvb38/).
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Corticoesteroides/efectos adversos , Calcio/uso terapéutico , Metaanálisis como Asunto , Osteoporosis/inducido químicamente , Osteoporosis/tratamiento farmacológico , Proyectos de Investigación , Revisiones Sistemáticas como Asunto/métodos , Vitamina D/uso terapéutico , Adulto , Humanos , Resultado del TratamientoRESUMEN
As one common kind of osteoporosis, postmenopausal osteoporosis (PMOP) is associated with the death and excessive loss of osteocytes. Estrogen deficiency of PMOP can cause osteocyte death by regulating necroptosis and apoptosis, but their roles in POMP have not been compared. In the present study, ovariectomy (OVX)-induced rat and murine long bone osteocyte Y4 (MLO-Y4) cells were used to compare the influence of necroptosis and apoptosis on osteocyte death and bone loss. Benzyloxycarbonyl-Val-Ala-Asp (zVAD) and necrostatin-1 (Nec-1) were used to specifically block cell apoptosis and necroptosis, respectively. OVX rats and MLO-Y4 cells were divided into zVAD group, Nec-1 group, zVAD + Nec-1 group, vehicle, and control group. The tibial plateaus of the rat model were harvested at 8 weeks after OVX and were analyzed by micro-computed tomography, transmission electron microscopy (TEM), the transferase dUTP nick end labeling assay, and western blot. The death of MLO-Y4 was stimulated by TNF-α and was measured by flow cytometry and TEM. The results found that necroptosis and apoptosis were both responsible for the death and excessive loss of osteocytes, as well as bone loss in OVX-induced osteoporosis, and furthermore necroptosis may generate greater impact on the death of osteocytes than apoptosis. Necroptotic death of osteocytes was mainly regulated by the receptor-interacting protein kinase 3 signaling pathway. Collectively, inhibition of necroptosis may produce better efficacy in reducing osteocyte loss than that of apoptosis, and combined blockade of necroptosis and apoptosis provide new insights into preventing and treating PMOP.
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OBJECTIVE: To explore the temperature change at the testis surface, apoptosis of spermatogenous cells and the expression of the heat shock protein 70 (HSP70) after scrotal reconstruction with the skin flap. METHODS: We included 36 healthy New Zealand white rabbits, 24 males and 12 females, in this study, and equally randomized the males into an experimental and a control group. The scrotal of the experimental rabbits were excised and reconstructed with the hypogastric flap, while the controls were left untreated. At the end of the 8th week after surgery, 6 animals were randomly taken from each of the two groups for measurement of the testis surface temperature and testicular biopsy. The apoptosis of spermatogenous cells in the testis tissues was detected by HE staining, and the expression of HSP70 determined by immunohistochemistry and imaging analysis. The other 6 animals exempt from testicular biopsy in each of the experimental and control groups were mated with the female rabbits, and observed for fertility. RESULTS: At the end of the 8th week after scrotal reconstruction, the testicular surface temperature was (38.1 +/- 0.6) degrees C in the experimental group, significantly higher than (36.0 +/- 0.30) degrees C before surgery (P < 0.05), and the apoptosis index (AI) of the spermatogenous cells was (71.85 +/- 2.7) %, as compared with (7.73 +/- 4.95) % in the control group (P < 0.05). The expression of HSP70 was found mainly in the spermatogenous cells of the experimental group and in the spermatoblasts of the control. A total of 6.0 +/- 1.3 baby rabbits were born in the control group, but none in the experimental group (P < 0.05). CONCLUSION: The testicular surface temperature rises after scrotal reconstruction with the hypogastric flap, which increases the apoptosis of spermatogenic cells and causes infertility. HSP70 is involved in protecting spermatogenic cells from apoptosis after scrotal reconstruction.
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Apoptosis , Proteínas HSP70 de Choque Térmico/metabolismo , Escroto/cirugía , Espermátides , Animales , Femenino , Masculino , Conejos , Espermátides/citología , Espermátides/metabolismo , Colgajos QuirúrgicosRESUMEN
Our earlier studies proved that RIPK3-mediated necroptosis might be an important mode of renal tubular cell death in rats with chronic renal injury and the necroptotic cell death can be triggered by tumor necrosis factor-α (TNF-α) in vitro, but the triggering role of angiotensin II (AngII), which exerts notable effects on renal cells for the initiation and progression of renal tubulointerstitial fibrosis, is largely unknown. Here, we identified the presence of necroptotic cell death in the tubular cells of AngII-induced chronic renal injury and fibrosis mice and assessed the percentage of necroptotic renal tubular cell death with the disruption of this necroptosis by the addition of necrostatin-1 (Nec-1). Furthermore, the observation was further confirmed in HK-2 cells treated with AngII and RIPK1/3 or MLKL inhibitors. The detection of Fas and FasL proteins led us to investigate the contribution of the Fas/FasL signaling pathway to AngII-induced necroptosis. Disruption of FasL decreased the percentage of necroptotic cells, suggesting that Fas and FasL are likely key signal molecules in the necroptosis of HK-2 cells induced by AngII. Our data suggest that AngII exposure might trigger RIPK3-MLKL-mediated necroptosis in renal tubular epithelial cells by activating the Fas/FasL signaling pathway in vivo and in vitro.
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Angiotensina II/farmacología , Proteína Ligando Fas/metabolismo , Túbulos Renales/citología , Necroptosis/efectos de los fármacos , Proteínas Quinasas/metabolismo , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Receptor fas/metabolismo , Animales , Línea Celular , Fibrosis , Humanos , Túbulos Renales/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Transducción de Señal/efectos de los fármacosRESUMEN
Abnormal renin-angiotensin system (RAS) activation plays a critical role in the initiation and progression of chronic kidney disease (CKD) by directly mediating renal tubular cell apoptosis. Our previous study showed that necroptosis may play a more important role than apoptosis in mediating renal tubular cell loss in chronic renal injury rats, but the mechanism involved remains unknown. Here, we investigate whether blocking the angiotensin II type 1 receptor (AT1R) and/or angiotensin II type 2 receptor (AT2R) beneficially alleviates renal tubular cell necroptosis and chronic kidney injury. In an angiotensin II (Ang II)-induced renal injury mouse model, we found that blocking AT1R and AT2R effectively mitigates Ang II-induced increases in necroptotic tubular epithelial cell percentages, necroptosis-related RIP3 and MLKL protein expression, serum creatinine and blood urea nitrogen levels, and tubular damage scores. Furthermore, inhibition of AT1R and AT2R diminishes Ang II-induced necroptosis in HK-2 cells and the AT2 agonist CGP42112A increases the percentage of necroptotic HK-2 cells. In addition, the current study also demonstrates that Losartan and PD123319 effectively mitigated the Ang II-induced increases in Fas and FasL signaling molecule expression. Importantly, disruption of FasL significantly suppressed Ang II-induced increases in necroptotic HK-2 cell percentages, and necroptosis-related proteins. These results suggest that Fas and FasL, as subsequent signaling molecules of AT1R and AT2R, might involve in Ang II-induced necroptosis. Taken together, our results suggest that Ang II-induced necroptosis of renal tubular cell might be involved both AT1R and AT2R and the subsequent expression of Fas, FasL signaling. Thus, AT1R and AT2R might function as critical mediators.
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Células Epiteliales/patología , Fallo Renal Crónico/patología , Túbulos Renales/patología , Receptor de Angiotensina Tipo 1/metabolismo , Receptor de Angiotensina Tipo 2/metabolismo , Angiotensina II/administración & dosificación , Angiotensina II/toxicidad , Bloqueadores del Receptor Tipo 1 de Angiotensina II/administración & dosificación , Bloqueadores del Receptor Tipo 2 de Angiotensina II/administración & dosificación , Animales , Línea Celular , Modelos Animales de Enfermedad , Humanos , Imidazoles/administración & dosificación , Fallo Renal Crónico/inducido químicamente , Túbulos Renales/citología , Losartán/administración & dosificación , Masculino , Ratones , Necroptosis/efectos de los fármacos , Oligopéptidos/administración & dosificación , Piridinas/administración & dosificación , Receptor de Angiotensina Tipo 2/agonistas , Transducción de Señal/efectos de los fármacosRESUMEN
Osteocyte apoptosis has been reported to play a central role in bone remodeling. In addition to apoptosis, other mechanisms may be involved in osteocyte loss. This study aimed to investigate the effect of necroptosis on osteocytes in ovariectomized (OVX) rats. Ninety-six female Sprague-Dawley rats were randomly divided into an OVX group and a sham group. At 0, 4, 8 and 12 weeks after surgery, specimens from each group (n = 12 each) were harvested. Bone mineral density (BMD) and body weight were measured. Transmission electron microscopy (TEM) and micro-CT were used to observe the changes in cellular morphology and bone microarchitecture induced by estrogen deficiency. Osteocyte apoptosis and necroptosis were evaluated via TUNEL and immunofluorescence staining for active caspase-3. At 8 weeks after ovariectomy, a greater number of osteocytes with typical necrotic morphological features were TUNEL positive but negative for active caspase-3. Western blotting, quantitative real-time PCR and immunofluorescence assessments demonstrated that the levels of receptor-interacting serine/threonine protein kinase 1 (RIP1) and RIP3 in osteocytes were significantly increased at 8 weeks after ovariectomy. These data are the first to suggest that necroptosis accelerates osteocyte loss under conditions of estrogen deficiency-induced osteoporosis in OVX rats. These findings provide evidence of a potential mechanism through which osteocyte necroptosis is associated with postmenopausal osteoporosis.
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Osteocitos/patología , Osteoporosis Posmenopáusica/patología , Proteínas Serina-Treonina Quinasas/metabolismo , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Animales , Apoptosis , Densidad Ósea , Femenino , Humanos , Osteocitos/citología , Osteocitos/metabolismo , Osteoporosis Posmenopáusica/etiología , Osteoporosis Posmenopáusica/metabolismo , Ovariectomía/efectos adversos , Proteínas Serina-Treonina Quinasas/análisis , Ratas Sprague-Dawley , Proteína Serina-Treonina Quinasas de Interacción con Receptores/análisisRESUMEN
Tubulointerstitial fibrosis (TIF) is caused by the progressive loss of renal tubular cells and the consequent replacement of the extracellular matrix. The progressive depletion of renal tubular cells results from apoptosis and necroptosis; however, the relative significance of each of these cell death mechanisms at different stages during the progression of chronic kidney disease (CKD) remains unclear. We sought to explore the mechanisms of renal tubular cell death during the early and intermediate stages of chronic renal damage of subtotal nephrectomied (SNx) rats. The results of tissue histological assays indicated that the numbers of necrotic dying cells and apoptotic cells were significantly higher in kidney tissues derived from a rat model of CKD. In addition, there was a significant increase in necroptosis observed by transmission electron microscopy (TEM) and an increase in the proportion of TUNEL-positive cells in kidney tissues from SNx rats compared with control rats, and necrostatin-1 (Nec-1) could inhibit necroptosis and reduce the proportion of TUNEL-positive cells. More importantly, we observed a significant increase in the incidence of necroptosis compared with apoptosis by TEM in vivo and in vitro and a significant increase in the proportion of TUNEL-positive tubular epithelial cells that did not express caspase-3 compared with those expressing cleaved caspase-3 in vitro. Furthermore, treatment with Nec-1 and zVAD strongly reduced necroptosis- and apoptosis-mediated renal tubular cell death and decreased the levels of blood urea nitrogen and serum creatinine and tubular damage scores of SNx rats. These results suggest that necroptotic cell death plays a more significant role than apoptosis in mediating the loss of renal tubular cells in SNx rats and that effectively blocking both necroptosis and apoptosis improves renal function and tubular damage at early and intermediate stages of CKD.
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Apoptosis , Túbulos Renales/patología , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Insuficiencia Renal Crónica/metabolismo , Insuficiencia Renal Crónica/patología , Animales , Progresión de la Enfermedad , Masculino , Necrosis , Ratas , Ratas Sprague-DawleyRESUMEN
Estrogen (E2) deficiency has been associated with accelerated osteocyte apoptosis. Our previous study showed necroptosis accelerated the loss of osteocytes in E2 deficiency-induced osteoporosis in rats in addition to apoptosis, but the mechanism involved remains. Necroptosis is a caspase-independent form of programmed cell death. In the necroptosis pathway, receptor interaction proteins 1 and 3 (RIP1/3) play vital roles. Necrostatin-1 (Nec-1) has been confirmed to be a specific inhibitor of necroptosis. However, the effect of Nec-1 on postmenopausal osteoporosis remains ambiguous. The aim of this study was to investigate the effect of Nec-1 on osteocytes in ovariectomized (OVX) rats. We found that an increased number of necroptotic osteocytes was related to the production of tumor necrosis factor-alpha (TNF-α) in OVX rats. Treatment with Nec-1 significantly decreased RIP1 and RIP3 expression in OVX rats and inhibited osteocyte necroptosis induced by TNF-α in vitro. Both E2 and Nec-1 treatment markedly ameliorated trabecular bone deterioration. Nec-1 also significantly elevated the levels of bone formation markers and decreased bone resorption markers. These data suggest that the role of Nec-1 on alleviating bone loss might be associated with Nec-1 restraining TNF-α-induced osteocyte necroptosis in rats with E2 deficiency-induced osteoporosis. This process may represent a novel therapeutic strategy for the treatment of postmenopausal osteoporosis.
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Apoptosis/efectos de los fármacos , Imidazoles/farmacología , Indoles/farmacología , Osteocitos/metabolismo , Osteoporosis Posmenopáusica/tratamiento farmacológico , Osteoporosis Posmenopáusica/metabolismo , Animales , Modelos Animales de Enfermedad , Femenino , Humanos , Osteocitos/patología , Osteoporosis Posmenopáusica/patología , Ovariectomía , Ratas , Ratas Sprague-DawleyRESUMEN
Graphene and its derivatives have been receiving increasing attention regarding their application in bone tissue engineering because of their excellent characteristics, such as a vast specific surface area and excellent mechanical properties. In this study, graphene-reinforced nanohydroxyapatite/polyamide66 (nHA/PA66) bone screws were prepared. The results of scanning electron microscopy observation and X-ray diffraction data showed that both graphene and nHA had good dispersion in the PA66 matrix. In addition, the tensile strength and elastic modulus of the composites were significantly improved by 49.14% and 21.2%, respectively. The murine bone marrow mesenchymal stem cell line C3H10T1/2 exhibited better adhesion and proliferation in graphene reinforced nHA/PA66 composite material compared to the nHA/PA66 composites. The cells developed more pseudopods, with greater cell density and a more distinguishable cytoskeletal structure. These results were confirmed by fluorescent staining and cell viability assays. After C3H10T1/2 cells were cultured in osteogenic differentiation medium for 7 and 14 days, the bone differentiation-related gene expression, alkaline phosphatase, and osteocalcin were significantly increased in the cells cocultured with graphene reinforced nHA/PA66. This result demonstrated the bone-inducing characteristics of this composite material, a finding that was further supported by alizarin red staining results. In addition, graphene reinforced nHA/PA66 bone screws were implanted in canine femoral condyles, and postoperative histology revealed no obvious damage to the liver, spleen, kidneys, brain, or other major organs. The bone tissue around the implant grew well and was directly connected to the implant. The soft tissues showed no obvious inflammatory reaction, which demonstrated the good biocompatibility of the screws. These observations indicate that graphene-reinforced nHA/PA66 composites have great potential for application in bone tissue engineering.
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Materiales Biocompatibles/farmacología , Durapatita/farmacología , Grafito/farmacología , Nanopartículas/química , Nylons/farmacología , Osteogénesis/efectos de los fármacos , Prótesis e Implantes , Fosfatasa Alcalina/genética , Fosfatasa Alcalina/metabolismo , Animales , Adhesión Celular/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Línea Celular , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Perros , Durapatita/química , Ratones , Nanopartículas/ultraestructura , Nylons/química , Osteocalcina/genética , Osteocalcina/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Difracción de Rayos XRESUMEN
OBJECTIVE: To explore the effect of burying testis in inguinal pocket on spermatogenesis. METHODS: Sixty New Zealand rabbits of 6-8 months old included 36 males and 24 females, weighing 2.5-2.7 kg. The male rabbits were randomly divided into the experimental group (n=18) and the control group (n=18). The model of repairing skin defect of scrotum were established by burying testes in inguinal region subcutaneously in the experimental group. The rabbits were not treated in the control group. The sperms were collected and the surface temperature of testis was measured in both groups after 8 weeks. Testes biopsies were harvested from 6 rabbits of 2 groups randomly respectively. The apoptosis of spermatogenic cells was detected with TUNEL. The other 12 male rabbits in two groups were fed respectively with female rabbits to observe the fertility. RESULTS: The semen density and the spermid activity ratio were (237.3 +/- 39.7) x 10(9)/L and 76.9% +/- 3.8% in the control group, and were (4.7 +/- 2.7) x 10(9)/L and 0 in the experimental group respectively; showing statistically significant difference between two groups (P< 0.05). The average superficial temperature of testes was (38.02 +/- 0.36) degrees C in the experimental group and (36.15 +/- 0.64) degrees C in the control group (P < 0.05). TUNEL results showed: The spermatogenic epithelium became thin and obvious apoptotic spermatogenic cells were found in experimental group; the spermatogenic epithelium was normal and few apoptotic spermatogenic cells were found in the control group. The apoptotic index (A1) was 89.69% +/- 3.76% in the experimental group and 7.73% +/- 4.95% in the control group (P < 0.05). The Pairing results showed that the female rabbits pairing with male rabbits of the experimental group were all not pregnant, and those of the control group were all pregnant (P < 0.05). CONCLUSION: As the same as the scrotum was reconstructed with skin flaps, it will induce the rabbit infertility that the testes were buried in inguinal region subcutaneously to repair defect of scrotum skin. The main reason is the excessive apoptosis of spermatogenic cell by the high testes environmental temperature.
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Apoptosis , Epitelio Seminífero/citología , Espermatogénesis , Testículo/cirugía , Animales , Femenino , Masculino , Embarazo , Conejos , Escroto/patologíaRESUMEN
OBJECTIVE: To explore the apoptosis and the express of proliferating cell nuclear antigen (PCNA) in spermatogenic cells, and study generation function of the rabbit after scrotal reconstruction with flaps. METHODS: The 48 New Zealand white rabbits were randomly divided into three groups as experimental group (the scrotum reconstructed with flaps, n = 48), the control group (the sham operated group, n = 24) and the blank group (n = 18). The apoptosis and the expression of PCNA in the spermatogenic cells were detected with TUNEL and the immunohistochemistry from the 3rd to the 8th week after operation. 8 weeks later, 12 animals in each group were fed respectively with one female rabbit to observe the procreation. RESULTS: The apoptotic index of the spermatogenic cells in blank group was 7.73 +/- 4.95. 3 weeks after operation, the apoptotic index of spermatogenic cells was 22.59 +/- 3.04 in the experimental group, and 21.13 +/- 1.68 in control group, showing no significant difference between the two groups (P > 0.05). 8 weeks after operation, the apoptotic index of spermatogenic cells was 71.85 +/- 2.69 in the experimental group, and 13.64 +/- 2.09 in control group, show a significant difference between them (P < 0.05). The apoptotic index in experimental group increased gradually from the 3rd to 8th week after scrotal reconstruction , which was markedly higher than that in the blank group (P < 0.05). The apoptotic index in control group was higher than that in the blank group at the 3rd week (P < 0.05), but not at the 8th week (P > 0.05). The proliferation index of spermatogenic cells was 9.32 +/- 9.30 and 12.52 +/- 3.87 in experimental group at the 3rd and 4th week, respectively, which was significantly lower than that in blank group (43.07 +/- 2.25) and control group (45.69 +/- 4.98) at the 3rd week (P < 0.05). The proliferation index of spermatogenic cells was 46.98 +/- 18.92 and 49.53 +/- 9.79 in experimental group at the 7th and 8th week, respectively, 39.90 +/- 5.10 in control group at the 8th week, showing no difference between the two groups (P > 0.05). The proliferation index of spermatogenic cells in control group at the 3rd and 8th week was not different from that in the blank group (P > 0.05). The female pairing rabbits in the blank and control group were all pregnant, and the average childbirths were 6.0 +/- 1.28 and 5.92 +/- 1.31 respectively, with no difference between the two groups (P > 0.05). All the female pairing rabbits in the experimental group were not pregnant, showing a significant difference from those in the blank and control group (P < 0.05). CONCLUSIONS: The rabbit generation functional disturbance after scrotal reconstruction with flaps is due to the excessive apoptosis of spermatogenic cell. The spermatogenic cell proliferation is affected only in the early postoperative period, but can recover later.