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OBJECTIVE@#To investigate the role and molecular mechanism of exosomal miR-224-5p in colorectal cancer (CRC).@*METHODS@#The miR-224-5p expression in CRC patient tissues and cell-derived exosomes was measured by laser capture microdissection and qRT-PCR, respectively. Dual-luciferase reporter gene assay was used to determine the target gene of miR-224-5p. The protein expressions of p53 and unc-51 like kinase 2 (ULK2) in CRC cells were detected by western blot. Flow cytometry was used to detect cell cycle and apoptosis. Cell proliferation was measured by CCK8 and EdU assay.@*RESULTS@#The miR-224-5p expression was upregulated in CRC tissues and increased progressively with the rise of CRC stage. CRC cells secreted extracellular miR-224-5p mainly in an exosome-dependent manner, and then miR-224-5p could be transferred to surrounding tumor cells to regulate cell proliferation in the form of autocrine or paracrine. Moreover, ULK2 was characterized as a direct target of miR-224-5p and was downregulated in CRC tissues. Interestingly, ULK2 inhibited CRC cell proliferation in a p53-dependent manner. Furthermore, exosome-derived miR-224-5p partially reversed the proliferation regulation of ULK2 on CRC cells.@*CONCLUSION@#Our findings demonstrate that exosome-transmitted miR-224-5p promotes p53-dependent cell proliferation by targeting ULK2 in CRC, which may offer promising targets for CRC prevention and therapy.
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Humanos , MicroARNs/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Exosomas/metabolismo , Proliferación Celular/genética , Neoplasias Colorrectales/genética , Línea Celular Tumoral , Regulación Neoplásica de la Expresión GénicaRESUMEN
Objective@#To investigate the prevalence and associated factors of unaided visual impairment and myopia among primary and secondary school students in Yunnan Province, and to provide scientific basis for myopia prevention.@*Methods@#The study was conducted among primary and secondary school students in Mangshi, Yunnan Province from March to August, 2014. All the 7 681 subjects underwent detailed eye examinations and a questionnaire survey. Myopia was defined as spherical equivalent value of less than -0.5 diopters. Unaided visual impairment was analyzed on the basis of the better and the worse-seeing eye, respectively.@*Results@#The prevalence of myopia and high myopia were 39.1% and 0.6%. The prevalence of unaided visual impairment was 11.4% based on the worse-seeing eye. Refractive errors accounted for 87.3% of the participants with unaided visual impairment. Prevalence of myopia was higher in girls than in boys (χ2=29.74, P<0.01), but there was no gender difference in high myopia (P=0.19). The prevalence of myopia and high myopia increased significantly with increasing age (χ2=351.23, 22.56, P<0.01). Besides, prevalence of myopia was 63.7% in Dai nationality students and 36.6% in Yi nationality students (χ2=78.14, P<0.01), which was higher than other ethnic minorities. After adjusting for the effects of sex, age and ethnicity, the presence of myopia was associated with increasing height (OR=1.02, 95%CI=1.01-1.03), computer use (OR=1.17, 95%CI=1.03-1.32), having a myopic father (OR=1.56, 95%CI=1.24-1.94), having a myopic mother (OR=1.33, 95%CI=1.08-1.63) and more time reading(OR=1.18, 95%CI=1.09-1.28). High myopia was found to be more prevalent in children who had a myopic father (OR=3.98, 95%CI=1.72-9.22) and using computers (OR=2.31, 95%CI=1.17-4.57).@*Conclusion@#Myopia and unaided visual impairment is prevalent in school students in rural China (Yunnan), though the prevalence is relatively lower compared with other areas in China. Attention should be paid to the formulation and input of primary eye care policies.
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<p><b>OBJECTIVE</b>To study the expression of neonatal Fc receptor in podocytes in human nephritis and immune-induced rat nephritis models: anti-Thy1.1 nephritis and Heymann nephritis.</p><p><b>METHODS</b>Thirty-nine cases of renal biopsies were enrolled from September 2009 to February 2010, including 8 cases of minimal change disease, 4 cases of focal segmental glomerulosclerosis, 9 cases of membranous nephropathy, 12 cases of IgA nephropathy and 6 cases of lupus nephritis. Five normal kidney tissue samples adjacent to renal clear-cell carcinoma were served as normal controls. Laser capture microdissection and real-time RT-PCR were used to assess the expression level of FcRn mRNA in glomeruli of various glomerulonephritides, and immunohistochemistry (IHC) of FcRn by SuperVision method was performed. In addition, rat models of mesangial proliferative nephritis (anti-Thy1.1 nephritis) and passive membranous nephropathy (Heymann nephritis) were established and FcRn was examined in renal tissues by IHC.</p><p><b>RESULTS</b>The FcRn mRNA level in lupus nephritis was statistically higher than that of normal controls (P < 0.05). FcRn protein expression by IHC was seen in lupus nephritis (6/6), membranous nephropathy (6/9) and IgA nephropathy (7/12), significantly higher than that of normal controls (0/5), P < 0.05. Minimal change disease and focal segmental glomerular sclerosis showed minimal or none expression of FcRn (1/8, 0/4 respectively) and not statistically difference from that of normal controls. Furthermore, FcRn expression in podocytes was detected in rat anti-Thy1.1 (3/5) and Heymann nephritis models (2/7) but was not detected in normal controls.</p><p><b>CONCLUSIONS</b>Expression of FcRn in podocytes was up-regulated in immune-induced human nephritis and rat nephritis models of anti-Thy1.1 nephritis and Heymann nephritis. FcRn may play a role in the development of immune-induced glomerulonephritis.</p>
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Animales , Humanos , Masculino , Ratas , Glomerulonefritis por IGA , Metabolismo , Patología , Glomerulonefritis Membranosa , Metabolismo , Patología , Glomeruloesclerosis Focal y Segmentaria , Metabolismo , Patología , Antígenos de Histocompatibilidad Clase I , Genética , Metabolismo , Captura por Microdisección con Láser , Nefritis Lúpica , Metabolismo , Patología , Nefritis , Genética , Alergia e Inmunología , Metabolismo , Patología , Nefrosis Lipoidea , Metabolismo , Patología , Podocitos , Metabolismo , ARN Mensajero , Metabolismo , Ratas Sprague-Dawley , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores Fc , Genética , Metabolismo , Antígenos Thy-1 , Alergia e Inmunología , Metabolismo , Regulación hacia ArribaRESUMEN
<p><b>OBJECTIVE</b>To study the biological impact and mechanism of recombinant tissue factor pathway inhibitor (rTFPI) on apoptosis of rat kidney mesangial cells (MsC).</p><p><b>METHODS</b>TFPI expression in human glomerular minor lesion (GML), mesangial proliferative glomerulonephritis (MPGN) and cultured rat MsC was detected using immunohistochemistry and immunofluorescence, respectively. Rat MsC were incubated with rTFPI and its variant peptides. Morphological changes of apoptosis were investigated by Hoechst 33258 and the apoptotic rate was assessed by flow cytometry. DNA fragmentation and effect of rTFPI on expression of caspase-3, Fas and bcl-2 were studied using gel electrophoresis and Western blot respectively.</p><p><b>RESULTS</b>The expression of TFPI in MPGN was higher than that in GML. TFPI was expressed in cultured rat mesangial cells. Apoptosis of MsC was induced by rTFPI, especially by its C-termianl, in a dose- and time-dependent manner. Apoptosis ratios of MsC treated with rTFPI were 2.1, 3.0 and 4.9 times more than control, respectively. Expression of gene caspase-3 and Fas was up-regulated in a dose-dependent manner wherease bcl-2 expression did not show any changes.</p><p><b>CONCLUSION</b>rTFPI induces apoptosis in cultured rat mesangial cells by its C-terminal possibly via Fas/FasL pathway.</p>
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Animales , Humanos , Ratas , Apoptosis , Fisiología , Caspasa 3 , Metabolismo , Células Cultivadas , Fragmentación del ADN , Citometría de Flujo , Lipoproteínas , Metabolismo , Farmacología , Células Mesangiales , Biología Celular , Metabolismo , Péptidos , Farmacología , Proteína Letal Asociada a bcl , Metabolismo , FarmacologíaRESUMEN
<p><b>OBJECTIVE</b>To study the clinicopathologic features of microscopic polyangitis (MPA), and to compare the differences in anti-neutrophil cytoplasmic autoantibody (ANCA)-positive and ANCA-negative patients, as well as in ANCA-positive cases with or without glomerular immunoglobulin deposition.</p><p><b>METHODS</b>Thirty-four biopsy-proven cases of MPA were retrieved from the archival files of the Department during the past 7 years. The clinicopathologic characteristics between ANCA-positive and negative patients, as well as between ANCA-positive cases with and without glomerular immunoglobulin deposition, were compared.</p><p><b>RESULTS</b>Amongst the 34 MPA patients studied, about one-fifth to one-half were accompanied by various extrarenal symptoms. Serum ANCA was positive in 26 patients (76.5%). A slight to moderate increase in urinary protein was demonstrated in 31 patients, while 3 patients had nephrotic syndrome. Elevated serum creatinine was detected in 32 cases. Renal biopsy revealed crescentic glomerulonephritis in 24 cases, focal segmental glomerulonephritis in 8 cases, vascular fibrinoid necrosis with inflammation in 7 cases, intimal thickening of arterioles in 24 cases, interstitial inflammatory cells, including neutrophil infiltration (21 cases), in 29 cases. Crescentic formation was more common in the ANCA-positive group than in the ANCA-negative group (P < 0.05). Amongst the 26 ANCA-positive cases, 10 had glomerular immunoglobulin deposits (including 1 case with IgA nephropathy). In general, these cases had a greater degree of proteinuria than those without glomerular immunoglobulin deposits (P < 0.05).</p><p><b>CONCLUSIONS</b>The diagnosis of MPA relies on histologic examination of renal biopsy and clinicopathologic correlation. Serum ANCA seems important for glomerular crescent formation. Glomerular immunoglobulin deposition may also play a significant role in the exacerbation of proteinuria.</p>
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Adulto , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Anticuerpos Anticitoplasma de Neutrófilos , Metabolismo , Biomarcadores , Biopsia , Glomerulonefritis , Metabolismo , Patología , Isotipos de Inmunoglobulinas , Metabolismo , Riñón , Patología , Enfermedades Renales , Metabolismo , Patología , Síndrome Nefrótico , Metabolismo , Patología , Proteinuria , Patología , Estudios Retrospectivos , Vasculitis , Metabolismo , PatologíaRESUMEN
<p><b>OBJECTIVE</b>To study the role of connective tissue growth factor (CTGF) in the development of glomerulosclerosis by experimental alteration of fibronectin (FN) and Type IV collagen (Col IV) expression in cultured rat mesangial cells (MsC).</p><p><b>METHODS</b>CTGF expression vector was transfected into MsC by Lipofectimine method. Protein and mRNA expression levels of CTGF, FN and Col IV were studied by Western blot and reverse transcription-polymerase chain reaction (RT-PCR) respectively.</p><p><b>RESULTS</b>Two of MsC clones (MCT-1 and MCT-2) with CTGF overexpression were successfully established and found to have significant increases of FN and Col IV at both protein and mRNA levels. Compared with the controls, the expression of FN protein and mRNA in the two clones were 3.2 times (P < 0.05) and 2.9 times (P < 0.05) higher respectively. The expression of Col IV protein and mRNA was 3.8 times (P < 0.01) and 2.4 times (P < 0.01) higher respectively.</p><p><b>CONCLUSION</b>CTGF up-regulates FN and Col IV expression in MsC and may play an important role in the development of glomerulosclerosis.</p>
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Animales , Ratas , Western Blotting , Células Cultivadas , Colágeno Tipo IV , Genética , Metabolismo , Factor de Crecimiento del Tejido Conjuntivo , Genética , Metabolismo , Fibronectinas , Genética , Metabolismo , Vectores Genéticos , Genética , Células Mesangiales , Biología Celular , Metabolismo , ARN Mensajero , Genética , Metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , TransfecciónRESUMEN
<p><b>BACKGROUND</b>Adrenomedullin (ADM), a potent hypotensive small peptide, was recently found to inhibit the proliferation of glomerular mesangial cells (MsC) in vitro and to attenuate glomerular lesions in vivo, however the mechanisms remain poorly understood. In this study, we attempted to elucidate them using molecular signal transduction.</p><p><b>METHODS</b>Cultured rat MsC were treated with ADM and several inhibitors of signalling molecules. Methyl thiazoleterazolium (MTT) assay and BrdU incorporation method were employed for examining MsC proliferation. Western blot analysis was used for detecting total mitogen activated protein kinases (t-MAPKs) and phosphorylated MAPKs (p-MAPKs) proteins.</p><p><b>RESULTS</b>ADM suppressed MsC proliferation in a concentration- and time-dependent fashion. This response was inhibited by ADM receptor antagonist CGRP8-37 and a potent protein kinase-A (PKA) inhibitor, H89. Forskolin, a direct adenylate cyclase activator, also significantly inhibited MsC proliferation. SB203580, a P38MAPK inhibitor, and U0126, a MEK inhibitor, both completely blocked ADM mediated responses in MsC. However, curcumin, a SAPK/JNK inhibitor, and GF109203X, a potent protein kinase-C (PKC) inhibitor, had no effect on MsC growth. Western blot analysis showed that ADM did not change the expression of t-MAPKs but increased p-SAPK/JNK and p-P38MAPK levels and decreased p-ERK level. These responses were inhibited by CGRP8-37. All these kinase phosphorylations, except for the increase in p-SAPK/JNK, could be stimulated using forskolin. In addition, only ADM mediated changes in ERK and P38MAPK phosphorylations were inhibited by H89. GF109203X did not affect ADM induced changes in three p-MAPKs expressions.</p><p><b>CONCLUSIONS</b>ADM inhibits MsC proliferation possibly through cAMP-PKA pathway. Both phosphorylations of ERK and P38MAPK pathways were necessary in mediating the antiproliferative response of ADM. It does not preclude the involvement of cAMP independent pathways in the ADM mediated responses.</p>
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Animales , Ratas , Adrenomedulina , Proliferación Celular , Células Cultivadas , Quinasas MAP Reguladas por Señal Extracelular , Fisiología , Mesangio Glomerular , Biología Celular , Proteínas Quinasas JNK Activadas por Mitógenos , Fisiología , Péptidos , Farmacología , Transducción de Señal , Fisiología , Proteínas Quinasas p38 Activadas por Mitógenos , FisiologíaRESUMEN
<p><b>OBJECTIVE</b>To study the effect of aldose reductase (AR) on expression of fibronectin and collagen IV in cultured rat renal mesangial cells (MsC).</p><p><b>METHODS</b>AR expression plasmid vector (pCDNA3-AR) was constructed by restriction endonuclease digestion and ligation procedures. Stable expression of AR in MsC was established by Lipofectin transfection. Western blot and immunofluorescence analyses were performed to verify the transfection efficiency. Expression of fibronectin and collagen IV proteins were analyzed using Western blot.</p><p><b>RESULTS</b>Expression of fibronectin and collagen IV in naive MsC treated with TGF-beta1 was upregulated in comparison to that of the untreated naive MsC (P < 0.01). MsC transfected with pCDNA3-AR showed a remarkable increase of expression of fibronectin and collagen IV (P < 0.01). Aldose reductase inhibitors (Sorbinil and Zopolrestat) significantly inhibited the expression of fibronectin and collagen IV in naive MsC (P < 0.05).</p><p><b>CONCLUSIONS</b>Overexpression or inhibition of AR activity significantly alters the expression of fibronectin and collagen IV proteins in cultured rat MsC, suggesting that AR plays a significant role in the pathogenesis of glomerulosclersis.</p>
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Animales , Ratas , Aldehído Reductasa , Genética , Metabolismo , Benzotiazoles , Farmacología , Células Cultivadas , Colágeno Tipo IV , Metabolismo , Fibronectinas , Metabolismo , Vectores Genéticos , Imidazolidinas , Farmacología , Células Mesangiales , Metabolismo , Ftalazinas , Farmacología , Plásmidos , Proteínas Recombinantes , Genética , Metabolismo , Transfección , Factor de Crecimiento Transformador beta1 , FarmacologíaRESUMEN
<p><b>OBJECTIVE</b>To study the changes of fibronectin (FN) and type IV collagen (ColIV) expression in cultured rat mesangial cells (MsC) transfected with Smad 2 vector and to investigate the molecular mechanism of glomerular extracellular matrix accumulation in glomerulosclerosis via transforming growth factor-beta (TGF-beta)/Smad signal pathway.</p><p><b>METHODS</b>Smad 2 vector was transfected into MsC by calcium phosphate. Western blot analysis was used to detect Smad 2 protein. The expression of FN and ColIV proteins and their mRNAs was determined by Western blot and reverse transcriptase-polymerase chain reaction respectively.</p><p><b>RESULTS</b>Four MsC clones (T-12, T-31, T-35, T-40) with Smad 2 overexpression were established. The expression of FN and ColIV was significantly increased at mRNA and protein levels in two (T-12, T-31). Compared with controls, the expression of FN proteins and mRNAs in these two clones was 2.4 times (P < 0.05) and 2.7 times (P < 0.05) higher respectively. The expression of ColIV proteins and mRNAs was 2.9 times (P < 0.01) and 3.3 times (P < 0.01) higher respectively.</p><p><b>CONCLUSIONS</b>It is postulated that Smad 2 in TGF-beta/Smad signal pathway is important in promoting the accumulation of FN and ColIV in sclerotic glomeruli of diseased kidneys.</p>