Immunolocalization of the TGFB1 system in submandibular gland fibrosis after experimental periodontitis in rats

Saliva is the first barrier to entry of bacteria and viruses into the body. The submandibular glands (SMG) contribute to the maintenance of oral health and regulation of immune/ inflam matory responses. Previous studies suggest that transforming growth factor beta 1 (TGFB1) may contribute to salivary gland fibrosis but the expression of the TGFB1 system in the SMG has not been elucidated. Thus, the aim of this study was to analyze in rat SMG the immunolocalization of TGFB1 and its specific receptors ALK5 (profibrotic) and ALK1 (proproliferative) and the coreceptor endoglin (EDG) in a bilateral experimental periodontitis (EP) model (cotton thread ligature around the neck of the first lower molars) for 1 and 6 weeks. Fixed SMG were embedded in paraffin and serially cut for routine hematoxylin-eosin staining for histological analysis or immunohistochemical techniques by diaminobenzidine detection. SMG histology from animals with EP showed timedependent structural changes involving marked reduction in the height of the contoured ducts, cell destruction, loss of secretory granules, periductal congestion and excess connective tissue surrounding these ducts indicative of a fibrotic process, compared to control SMG. TGFB1, ALK5 and ALK1 receptors and the coreceptor EDG were mainly immunolocalized in ductal cells and in the fibrotic areas in EP groups. The expression of the profibrotic ALK5 receptor was increased in areas of fibrosis in SMG of animals with EP. In SMG of rats with EP, the localization of the TGFB1 specific receptors in the ducts and cells from fibrotic areas, due to the expression of TGFB1 in the surrounding areas, might indicate paracrine and autocrine actions exerted by TGFB1 via its specific receptors. The results of this study suggest that TGFB1 promotes fibrosis, inducing cell proliferation via ALK1 and EDG receptors and stimulates fibrosis relatedprocesses via ALK5 receptor, which could lead to abnormal secretor activity of the SMG during periodontal disease.

La saliva es la primera barrera para la entrada de bacterias y virus en el cuerpo. Las glándulas submandibulares (GSM) contribuyen al mantenimiento de la salud oral y a la regulación de las respuestas inmunoinflamatorias. Estudios previos sugieren que el factor de crecimiento transformante beta 1 (TGFB1) puede contribuir a la fibrosis de las glándulas salivales, pero la expresión y localización del sistema TGFB1 en las GSM no ha sido dilucidada. El objetivo del presente trabajo fue analizar por inmunohistoquímica en las GSM de ratas la expresión de TGFB1 y sus receptores específicos ALK5 (profibrótico) y ALK1 (proproliferativo) y el coreceptor endoglina (EDG) en un modelo de periodontitis bilateral experimental (PE) (hilo de algodón alrededor del cuello de los primeros molares inferiores) durante 1 y 6 semanas. Las GSM fueron fijadas y embebidas en parafina para realizar cortes seriados los cuales se tiñeron con hematoxilinaeosina para analizar la histología o se procesaron para realizar la técnica de inmunohistoquímica mediante detección con diaminobenzidine. La histología de las GSM de animales con PE reveló cambios estructurales tiempo dependientes, con una marcada reducción de la altura de los conductos, destrucción celular, pérdida de gránulos secretores, congestión periductal y exceso de tejido conectivo que rodea los conductos, indicando un proceso de fibrosis respecto de las GSM de animales control. TGFB1, ALK5 y ALK1 y el coreceptor EDG fueron principalmente inmunolocalizados en las células que forman los ductos y en las áreas de fibrosis en los grupos con PE. La expresión del receptor profibrótico ALK5 se incrementó en las áreas de fibrosis en GSM de animales con PE. En GSM de ratas con PE, la localización de los receptores específicos de TGFB1 en las células de los conductos y áreas de fibrosis, junto con la expresión de TGFB1 en las áreas circundantes, podría indicar acciones paracrinas y autocrinas ejercidas por TGFB1 a través de sus receptores específicos. Los resultados de este estudio sugieren que TGFB1 podría inducir un proceso de fibrosis promoviendo la proliferación celular a través de los receptores ALK1 y EDG, y favoreciendo procesos relacionados con la fibrosis a través de su receptor ALK5, lo que conduciría a una actividad secretora anormal de la GSM durante la enfermedad periodontal.

Differences in the expression of inhibin receptors and activin receptors in normal human ovaries and their significance / 中华妇产科杂志

Objective To explore the differences in the expression of inhibin(INH)receptors and activin (ACT)receptors in the follicular/luteinic phase in normal human ovaries and their relationship with female endocrine hormone levels.Methods Real time PCR and immunohistochemistry were used to determine the expression of inhibin receptors(INHR)genes,activin receptors(ACTR)genes.Serum estradiol(E2),follicle stimulating hormone(FSH),luteinizing hormone(LH),INHB,ACTA levels were determined by a solid quantitative sandwich enzyme immunoassay technique(Sandwich ELISA)in 21 women during follicular phase and another 21 women during luteinic phase,the correlations between each gene and each hormone were analyzed.Results(1)ACT type Ⅰ and Ⅱ receptors genes(ACTR Ⅰ A,ACTR Ⅰ B,ACTRⅡA,ACTR Ⅱ B)and INH receptor β-glycan genes were expressed higher in the follicular phase than in the luteinic phase:ACTR Ⅰ A(0.50±0.17 vs 0.36±0.18;P＜0.05),ACTR Ⅰ B(0.050±0.019 vs0.036±0.020;P＜0.05),ACTRⅡ A(0.10±0.04 vs 0.07±0.04;P＜0.05),ACTR Ⅱ B(0.28±0.10vs 0.19±0.11;P＜0.05),β-glycan(0.26±0.10 vs 0.17±0.09;P＜0.01).(2)The intensities of ACTR I A,ACTR Ⅱ A,β-glycan immunostaining in human normal ovaries in the follicular phase were significantly stronger compared to those in luteinic phase.In the follicular phase β-glycan expression was positively correlated with serum E2,FSH,LH,INHB levels.The correlation coefficient was 0.53(P＜0.05).0.74(P＜0.01),0.85(P＜0.01)and 0.76(P＜0.01)respectively.Conclusion In normal human ovary in the follicular phase INH and ACT bind their receptors and down-regulate or up-regulate FSH,thus influencing the follicular development.

The activin receptor-like kinase Ⅰ promotes proliferation and migration in HUVECs / 中国病理生理杂志

AIM: To study the function of activin receptor-like kinase Ⅰ(ALK1) gene in vascular endothelium.METHODS: The human umbilical vein endothelial cells(HUVECs) were cultured,and the change of expression of ALK1,ALK5 in activation of HUVECs was analyzed.The full-leng coding sequence of ALK1 was cloned into pcDNA3.1+using standard protocols.The constructed pcDNA3.1+ALK1 plasmid were transfected into HUVECs.The proliferation and migration of HUVECs were detected by boyden champer and flow cytometry.RESULTS: The expression of ALK1 was up-regulated in resolution.ALK1 promoted the proliferation and migration of HUVECs.CONCLUSION: ALK1 has an important function in remodeling by promoting the proliferation and migration of endothelial cells.