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Objective:To determine the expression of silent information regulator 1 (Sirt1) , Sirt3 and hypoxia-inducible factor 1α (HIF-1α) in cutaneous squamous cell carcinoma (CSCC) tissues and cells, and to explore their role in the occurrence and development of CSCC.Methods:From January 2019 to December 2020, 30 lesional skin tissues were obtained from patients with histopathologically confirmed poorly-, moderately- or well-differentiated CSCC, and 30 normal skin tissues were obtained from patients with non-cancerous diseases in Department of Dermatology, General Hospital of Ningxia Medical University. A CSCC cell line A431 and a human keratinocyte cell line HaCaT were cultured. Immunohistochemical study, Western blot analysis and real-time quantitative PCR (RT-PCR) were performed to determine the protein and mRNA expression of Sirt1, Sirt3 and HIF-1α in CSCC tissues of different grades of differentiation and normal skin tissues, cytochemical and immunofluorescence staining and RT-PCR were conducted to determine the protein and mRNA expression of Sirt1, Sirt3 and HIF-1α in A431 and HaCaT cells, respectively. Comparisons of measurement data among multiple groups were performed by using one-way analysis of variance, and comparisons between two groups by using t test. Results:Immunohistochemical study showed that the expression level of Sirt3 (expressed as the average optical density) was 100 ± 12.12, 117.72 ± 26.23, 127.32 ± 24.45, 132.71 ± 31.61 in the normal skin tissues and well-, moderately- and poorly-differentiated CSCC tissues respectively, and there was a significant difference among these groups ( F = 20.14, P < 0.001) ; the expression of Sirt1 and HIF-1α increased in turn from the normal skin tissues to the well-, moderately- and poorly-differentiated CSCC tissues, and significantly differred in these groups ( F = 174.50, 225.00, respectively, both P < 0.001) . As Western blot analysis revealed, the expression level of Sirt3 significantly differed among the normal skin tissues, well-, moderately- and poorly-differentiated CSCC tissues (expressed as relative gray value: 1.000 ± 0.132, 1.403 ± 0.411, 1.387 ± 0.393, 1.677 ± 0.683, respectively; F = 34.97, P < 0.001) , and so did the expression levels of Sirt1 and HIF-1α ( F = 69.29, 199.90, respectively, both P < 0.00l) , with a gradually increasing trend in their expression levels from the the normal skin tissues to well-, moderately- and poorly-differentiated CSCC tissues. RT-PCR showed that the mRNA expression of Sirt3, Sirt1 and HIF-1α was sequentially increased from the normal skin tissues to well-, moderately- and poorly-differentiated CSCC tissues, and significant differences were observed among these groups ( F = 113.00, 174.50, 50.33, respectively, all P < 0.001) . The protein expression levels of Sirt3, Sirt1 and HIF-1α were significantly higher in the A431 cells than in the HaCaT cells ( t = 16.75, 18.34, 27.76, respectively, all P < 0.001) , and so were their mRNA expression levels ( t= 14.22, 9.62, 16.86, respectively, all P < 0.001) . Conclusion:Increased expression of Sirt3, Sirt1 and HIF-1α was observed in CSCC tissues and cells, which may promote the occurrence and development of CSCC.
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Objective To detect mutations in the ARAD1 gene in a pedigree with dyschromatosis symmetrica hereditaria (DSH).Methods Genomic DNA was extracted from the peripheral blood of 8 family members (including 5 patients with DSH and 3 unaffected members) in the pedigree with DSH,as well as 100 unrelated healthy controls.All the 15 exon sequences of the ADAR1 gene were amplified by polymerase chain reaction (PCR)followed by direct sequencing.Then,mutations were detected in comparison with the standard sequence of the ADAR1 gene in Genebank.Results A nonsense mutation C.1420C ＞ T (p.Arg474X) was identified at position 1 420 in exon 2 of the ADAR1 gene in the 5 patients with DSH,but not in the 3 unaffected members or 100 unrelated healthy controls.Conclusion The nonsense mutation C.1420C ＞ T in the ADAR1 gene is the causative mutation in the pedigree with DSH.
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Objective To investigate expressions of phosphorylated c-Jun N-terminal kinase (p-JNK)and P38 mitogen-activated protein kinase(p-P38MAPK)in psoriasis vulgaris lesions. Methods Tissue specimens were obtained from lesions of 30 patients with psoriasis vulgaris and normal skin of 30 healthy human controls. An immunohistochemical study and Western-blot analysis were performed to measure protein expressions of p-JNK and p-P38MAPK in these skin specimens. Results As the immunohistochemical study showed, the expressions of p-JNK and p-P38MAPK(expressed as the average optical density [AOD]value for targeted proteins)were significantly higher in psoriasis vulgaris lesions than in normal skin tissues (p-JNK: 0.663 ± 0.016 vs. 0.333 ± 0.009, t = 44.869, P < 0.001; p-P38MAPK: 0.436 ± 0.011 vs. 0.306 ± 0.010, t = 21.913, P < 0.001). Western-blot analysis also showed increased protein expressions of p-JNK and p-P38MAPK in psoriasis vulgaris lesions compared with normal skin tissues (t = 20.477, 165.084, respectively, both P <0.05). Conclusion The activation of JNK and P38MAPK may be involved in the overproliferation of epidermal cells in psoriasis vulgaris lesions.
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Background It is well known that diabetic retinopathy(DR) involves in complex pathogenesis.Worse outcome of brain ischemia with hyperglycemia is mediated by mitogen-activated protein kinases (MAPK).However,the effect of phosphorylation of MAPK signal pathway on DR needs further investigation.Objective This study was to explore the the possible role of extracellular signal-regulated kinase1/2(ERK1/2)in DR.Methods Sixty SPF SD rats aged 8-weeks were grouped into the control group and diabetes group.Diabetic models were established by intraperitoneal injection of streptozotocin (STZ) (60 mg/kg) dissolved in citric acid buffer.Blood glucose level was more than 16.7 mmol/L were used as the diabetic group.The equivalent amount of citric acid buffer was injected in the same way in the rats of the control group.The rats were sacrificed and retinas were isolated in 4 weeks and 12 weeks after modeling.The morphology of rat retinas was examined by hematoxylin and eosin staining.The relative expression levels of phosphorylated-ERK1/2 and glial fibrillary acidic protein (GFAP)(absorption,A value) in rat retinas were detected by immunohistochemistry.Results Blood glucose levels of rats were significantly higher than those in the diabetic group compared with the control group at both 4 and 12 weeks after modeling(t=14.174,13.771,both at P＜0.05).In addition,the body weight was significantly lower and the drinking was much more in the rats of the diabetic group in comparison with the control group in 12 weeks(t=8.670,18.725,both at P＜0.05).Twelve weeks after modeling,the decrease of retinal thickness,swelling of outer plexiform layer and decline of number of retinal ganglion cells,rods and cones were seen under the optical microscope.The relative expression levels of GFAP in the retinas were 3 197.1 ±13.1 and 7 202.0±56.8 in the diabetic group at the 4 and 12 weeks,which were significantly higher than those in the control group (2 152.8 ± 16.1 and 2 337.0±8.6) (t =6.327,16.417,both at P＜0.05).In 12 weeks after modeling,the relative expression level of phosphorylated-ERK 1/2 was significantly higher in the diabetic group compared with the control group (2 850.6±2.4 versus 1 274.6± 1.3),showing a significant difference between them (t =12.771,P ＜ 0.05).Conclusions Phosphorylation of ERK1/2 signal transduction pathway is involved in the STZ-induced DR through mediating the activation of Müller cells and inducing the apoptosis of photoreceptor and ganglione cells.
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Objective To investigate the expression and activation of extracellular signal-regulated kinase 1/2 (ERK 1/2), its upstream molecule, epidermal growth factor receptor (EGFR), and downstream transcription factor, Ets-like protein 1 (ELK-1), in lesions of psoriasis vulgaris, and to evaluate the relationship between ERK pathway and psoriasis vulgaris. Methods Tissue samples were obtained from the lesions of 40 patients with psoriasis vulgaris and normal skin of 20 normal human controls. Immunohistochemistry and Western blot were performed to detect the expressions of phosphorylated ERK1/2, EGFR and ELK-1 in the tissue samples.Results As immunohistochemistry showed, the integrated optical density (IOD) of p-ERK1/2, p-EGFR and p-ELK-1 was 269.85 ± 57.96, 136.88 ± 30.33 and 237.61 ± 56.29 respectively in the psoriatic lesions, significantly higher than that in the normal controls ( 140.24 ± 24.42, 110.66 ± 28.99 and 119.04 ± 21.99, respectively, all P ＜ 0.05). A positive correlation was observed between the expression of p-EGFR and p-ERK1/2(r = 0.57, P ＜ 0.05) and between that of p-ERK1/2 and p-ELK-1 (r=0.72,P＜0.05) in psoriatic lesions.Conclusion The enhanced signal transduction through phosphorylated EGFR→ERK1/2→ELK-1 pathway may play a certain role in the pathophysiological process of psoriasis vulgaris.