scholarly journals The Expression of Ptch1 and Ptch2 in the Stenotic Tissue of Congenital Ureteropelvic Junction Obstruction in Children

2020 ◽  
Author(s):  
Yanfang Yang ◽  
wenwen han ◽  
Weiping Zhang ◽  
Ning Sun
Keyword(s):  
Smooth Muscle ◽  
Western Blot ◽  
Smooth Muscle Cells ◽  
Real Time ◽  
Real Time Pcr ◽  
Ureteropelvic Junction Obstruction ◽  
Transmembrane Protein ◽  
Muscle Cells ◽  
Ureteropelvic Junction ◽  
Ureteral Polyps

Abstract Background: Ptch1 and Ptch2 are expressed in tubular epithelium and stromal cells adjacent to the UPJ. They mediate inhibition of Smoothened, a transmembrane protein expressed on the cell surface. If the pathway is disturbed, UPJOcan occur. This aim study aimed to determine the expression of Ptch1 (P1) and Ptch2 (P2) in stenotic segments in children with congenital ureteropelvic junction obstruction (UPJO) compared with normal control subjects. Methods: Stenotic segments of ureter tissues were obtained from 20 UPJO patients.UPJO caused by other pathogenies, such as vessel and ureteral polyps, were excluded. The control ureter specimens were obtained from 10 patients with Wilm’s tumor, and the tissues were confirmed histologically to be unaffected. Immunofluorescence, western blot and real-time PCR were used to investigate the expression of P1 and P2. Statistical methods were used to find the differences between the two groupsResults: P1 and P2 were identified in the cytoplasm of smooth muscle in two groups through immunohistochemistry. However, there were no statistical differences between the two groups in P1 and P2 with immunohistochemistry (P=0.31 and P=0.3, respectively). There were also no statistical differences with western blot (P=0.75 and P=0.9, respectively) and real-time PCR (P=0.52 and P=0.45, respectively). However, with the immunofluorescence it was found that red-stained P1 were diffused in the controls group, but were mainly located in the intracellular perinuclear compartment of smooth muscle cells in UPJO. Conclusions: The expression of P1 and P2 between the two groups had no statistical significant. P1 were mainly located in the intracellular perinuclear compartment of smooth muscle cells in UPJO. The P1 pathway might be disturbed by the abnormal distribution rather than the quantity, which might be one probable pathogenesis of UPJO.

scholarly journals The Expression of Ptch1 and Ptch2 in the Stenotic Tissue of Congenital Ureteropelvic Junction Obstruction in Children

2020 ◽  
Author(s):  
Yanfang Yang ◽  
wenwen han ◽  
Weiping Zhang ◽  
Ning Sun
Keyword(s):  
Smooth Muscle ◽  
Western Blot ◽  
Smooth Muscle Cells ◽  
Real Time ◽  
Real Time Pcr ◽  
Ureteropelvic Junction Obstruction ◽  
Transmembrane Protein ◽  
Muscle Cells ◽  
Ureteropelvic Junction ◽  
Abnormal Distribution

Abstract Objective Ptch1 and Ptch2 are expressed in tubular epithelium and stromal cells adjacent to the UPJ. They mediated inhibition of smoothened, a transmembrane protein expressed on the cell surface. If the pathway was disturbed, UPJO might be onset. The aim of this study was to determine the expression of Ptch1 (P1) and Ptch2 (P2) in stenotic segments in children with congenital ureteropelvic junction obstruction (UPJO) versus in normal control subjects. Materials and methods Stenotic segments of ureter tissues were obtained from 20 UPJO patients. Extrinsic stenosis, such as vessel and ureteral polyp, were excluded. The control ureter specimens were obtained from 10 patients with Wilm’s tumor, and the tissues were confirmed histologically to be unaffected. Immunofluorescence, Western blot and real-time PCR were used to investigate the expression of P1 and P2. Statistical methods were used to find the differences between the two groups Results P1 and P2 were identified that located in the cytoplasm of smooth muscle in two groups through Immunohistochemistry. However, there were no statistical differences between the two groups in P1 and P2 with immunohistochemistry (P=0.31 and P=0.3, respectively). Meanwhile there were no statistical differences with Western blot (P=0.75 and P=0.9, respectively) and real-time PCR (P=0.52 and P=0.45, respectively). But we found that in the immunofluorescence P1 were diffused in controls and mainly surrounding the nucleuses of smooth muscle cells in UPJO. Conclusions The expression of P1 and P2 between the two group had no statistically significant. P1 mainly surrounding the nucleuses of smooth muscle cells in UPJO. The P1 pathway might be disturbed by the abnormal distribution of P1 rather than the quantity.

Altered Cytoskeletal Structure of Smooth Muscle Cells in Ureteropelvic Junction Obstruction

2011 ◽  
Vol 185 (6) ◽  
pp. 2314-2319 ◽  
Author(s):  
Giuseppina Cutroneo ◽  
Salvatore Arena ◽  
Giuseppe Anastasi ◽  
Raimondo M. Cervellione ◽  
Silvia Grimaldi ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Ureteropelvic Junction Obstruction ◽  
Muscle Cells ◽  
Ureteropelvic Junction ◽  
Cytoskeletal Structure

1α,25-Dihydroxy-vitamin D3 stimulation of bronchial smooth muscle cells induces autocrine, contractility, and remodeling processes

2007 ◽  
Vol 29 (2) ◽  
pp. 161-168 ◽  
Author(s):  
Yohan Bossé ◽  
Karim Maghni ◽  
Thomas J. Hudson
Keyword(s):  
Vitamin D ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Real Time ◽  
Real Time Pcr ◽  
Muscle Cells ◽  
Fold Change ◽  
Bronchial Smooth Muscle ◽  
Bronchial Smooth Muscle Cells ◽  
Unique Genes

Genetic variants in the vitamin D receptor (VDR) gene were recently associated with asthma. The biological mechanisms explaining this association are unknown but are likely to involve many cell types given the pleiotropic effect of its ligand, 1α,25-dihydroxy-vitamin D3 [1α,25(OH)2D3]. Considering the prominent role of bronchial smooth muscle cells (BSMCs) in the pathogenesis of asthma, experiments were conducted to explore the gene regulatory effects of 1α,25(OH)2D3 in these cells. Using RT-PCR and Western blot, we showed that VDR is present both at the mRNA transcript and protein levels in human BSMCs. The functionality of the receptor was then demonstrated by showing a >200-fold change in the expression of the 24-hydroxylase (CYP24A1) gene following 1α,25(OH)2D3 stimulation. Microarray experiments were then performed to identify differentially regulated genes and pathways in BMSCs treated or not with 1α,25(OH)2D3. A total of 729 probe sets on the U133 plus 2.0 Affymetrix GeneChip showed fold-change differences above the 1.5 threshold using the Robust Multichip Average intensities. This corresponds to 231 unique genes that were upregulated and 215 unique genes that were down-regulated following 1α,25(OH)2D3 stimulation. A high similarity between microarray and real-time PCR results was observed for 13 random genes, with a concordance correlation coefficient of 0.91. Real-time PCR was also performed to confirm the regulation of asthma candidate genes. To identify the biological relevance of this regulation, biological pathways analyses were performed. The most significant network of upregulated genes included genes involved in morphogenesis, cell growth, and survival as well as genes encoding structural proteins, which are potentially involved in airway remodeling.

Altered Cytoskeletal Structure of Smooth Muscle Cells in Ureteropelvic Junction Obstruction

2009 ◽  
Vol 5 ◽  
pp. S26-S27
Author(s):  
Salvatore Arena ◽  
Angelo Favaloro ◽  
Maria Grazia Scuderi ◽  
Maria Teresa Sinatra ◽  
Noemi Cantone ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Ureteropelvic Junction Obstruction ◽  
Muscle Cells ◽  
Ureteropelvic Junction ◽  
Cytoskeletal Structure

Laser-assisted microdissection and real-time PCR detect anti-inflammatory effect of perfluorocarbon

2003 ◽  
Vol 285 (1) ◽  
pp. L55-L62 ◽  
Author(s):  
Katharina von der Hardt ◽  
Michael Andreas Kandler ◽  
Ludger Fink ◽  
Ellen Schoof ◽  
Jörg Dötsch ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Mrna Expression ◽  
Vascular Smooth Muscle Cells ◽  
Alveolar Macrophages ◽  
Real Time Pcr ◽  
Muscle Cells ◽  
Cell Types ◽  
Anti Inflammatory ◽  
Inflammatory Effect

The aim of this study was to identify cell types involved in the anti-inflammatory effect of ventilation with perfluorocarbon in vivo. Fifteen anesthetized, surfactant-depleted piglets received either aerosolized perfluorocarbon (Aerosol-PFC), partial liquid ventilation (rLV) at functional residual capacity (FRC) volume (FRC-PLV), or intermittent mandatory ventilation (control). After laser-assisted microdissection of different lung cell types, mRNA expression of IL-8 and ICAM-1 was determined using TaqMan real-time PCR normalized to hypoxanthine phosphoribosyltransferase (HPRT). IL-8 mRNA expression (means ± SE; control vs. Aerosol-PFC) was 356 ± 142 copies IL-8 mRNA/copy HPRT mRNA vs. 3.5 ± 1.8 in alveolar macrophages ( P <0.01); 208 ± 108 vs. 2.7 ± 0.8 in bronchiolar epithelial cells ( P <0.05); 26 ± 11 vs. 0.7 ± 0.2 in alveolar septum cells ( P <0.01); 2.8 ± 1.0 vs. 0.8 ± 0.4 in bronchiolar smooth muscle cells ( P <0.05); and 1.1 ± 0.4 vs. 0.2 ± 0.05 in vascular smooth muscle cells ( P <0.05). With FRC-PLV, IL-8/HPRT mRNA expression was significantly lower in macrophages, bronchiolar epithelial, and vascular smooth muscle cells. ICAM-1 mRNA expression in vascular endothelial cells remained unchanged. Predominantly, alveolar macrophages and bronchiolar epithelial cells were involved in the inflammatory pulmonary process. The anti-inflammatory effect of Aerosol-PFC was most pronounced.

scholarly journals Akt/eNOS and MAPK Signaling Pathways Mediated the Phenotypic Switching of Thoracic Aorta Vascular Smooth Muscle Cells in Aging/Hypertensive Rats

2018 ◽  
pp. 543-553 ◽  
Author(s):  
L. ZHANG ◽  
Z. XU ◽  
Y. WU ◽  
J. LIAO ◽  
F. ZENG ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Western Blot ◽  
Smooth Muscle Cells ◽  
Thoracic Aorta ◽  
Vascular Smooth Muscle Cells ◽  
Muscle Cells ◽  
Mapk Signaling ◽  
Phenotypic Switching ◽  
Hypertensive Rats

Considerable evidence demonstrates that phenotypic switching of vascular smooth muscle cells (VSMCs) is influenced by aging and hypertension. During phenotypic switching, VSMCs undergo a switch to a proliferative and migratory phenotype, with this switch being a common pathology in cardiovascular diseases. The aim of this study was to explore the joint influence of age and hypertension on thoracic aortic smooth muscle phenotypic switching and the balance of Akt and mitogen-activated protein kinase (MAPK) signaling during this switch. Different ages of spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY) were used to establish hypertension and aging models. The phenotypic state was determined by detecting the marker proteins α-SM-actin, calponin, and osteopontin (OPN) via immunohistochemical staining and Western blot. Signaling proteins associated with the Akt and MAPK pathways were detected in rat thoracic aorta using Western blot. Both aging and hypertension caused a decrease in contractile (differentiated) phenotype markers (α-SM-actin and calponin), while the synthetic (proliferative or de-differentiated) phenotype maker was elevated (OPN). When combining hypertension and aging, this effect was enhanced, with Akt signaling decreased, while MAPK signaling was increased. These results suggested that VSMCs phenotype switching is modulated by a balance between Akt and MAPK signaling in the process of aging and hypertension.

scholarly journals Role of 2′,3′-cyclic nucleotide 3′-phosphodiesterase in the renal 2′,3′-cAMP-adenosine pathway

2014 ◽  
Vol 307 (1) ◽  
pp. F14-F24 ◽  
Author(s):  
Edwin K. Jackson ◽  
Delbert G. Gillespie ◽  
Zaichuan Mi ◽  
Dongmei Cheng ◽  
Rashmi Bansal ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Real Time ◽  
Vascular Smooth Muscle Cells ◽  
Real Time Pcr ◽  
Cyclic Nucleotide ◽  
Kidney Tissue ◽  
Energy Depletion ◽  
Proximal Tubular

Energy depletion increases the renal production of 2′,3′-cAMP (a positional isomer of 3′,5′-cAMP that opens mitochondrial permeability transition pores) and 2′,3′-cAMP is converted to 2′-AMP and 3′-AMP, which in turn are metabolized to adenosine. Because the enzymes involved in this “2′,3′-cAMP-adenosine pathway” are unknown, we examined whether 2′,3′-cyclic nucleotide 3′-phosphodiesterase (CNPase) participates in the renal metabolism of 2′,3′-cAMP. Western blotting and real-time PCR demonstrated expression of CNPase in rat glomerular mesangial, preglomerular vascular smooth muscle and endothelial, proximal tubular, thick ascending limb and collecting duct cells. Real-time PCR established the expression of CNPase in human glomerular mesangial, proximal tubular and vascular smooth muscle cells; and the level of expression of CNPase was greater than that for phosphodiesterase 4 (major enzyme for the metabolism of 3′,5′-cAMP). Overexpression of CNPase in rat preglomerular vascular smooth muscle cells increased the metabolism of exogenous 2′,3′-cAMP to 2′-AMP. Infusions of 2′,3′-cAMP into isolated CNPase wild-type (+/+) kidneys increased renal venous 2′-AMP, and this response was diminished by 63% in CNPase knockout (−/−) kidneys, whereas the conversion of 3′,5′-cAMP to 5′-AMP was similar in CNPase +/+ vs. −/− kidneys. In CNPase +/+ kidneys, energy depletion (metabolic poisons) increased kidney tissue levels of adenosine and its metabolites (inosine, hypoxanthine, xanthine, and uric acid) without accumulation of 2′,3′-cAMP. In contrast, in CNPase −/− kidneys, energy depletion increased kidney tissue levels of 2′,3′-cAMP and abolished the increase in adenosine and its metabolites. In conclusion, kidneys express CNPase, and renal CNPase mediates in part the renal 2′,3′-cAMP-adenosine pathway.

Direct association and translocation of PKC-α with calponin

2004 ◽  
Vol 286 (6) ◽  
pp. G954-G963 ◽  
Author(s):  
Suresh B. Patil ◽  
Mercy D. Pawar ◽  
Khalil N. Bitar
Keyword(s):  
Smooth Muscle ◽  
Western Blot ◽  
Smooth Muscle Cells ◽  
Direct Interaction ◽  
Muscle Cells ◽  
Smooth Muscle Contraction ◽  
Direct Association ◽  
Translocated Proteins

Calponin has been implicated in the regulation of smooth muscle contraction through its interaction with F-actin and inhibition of the actin-activated MgATPase activity of phosphorylated myosin. Calponin has also been shown to interact with PKC. We have studied the interaction of calponin with PKC-α and with the low molecular weight heat-shock protein (HSP)27 in contraction of colonic smooth muscle cells. Particulate fractions from isolated smooth muscle cells were immunoprecipitated with antibodies to calponin and Western blot analyzed with antibodies to HSP27 and to PKC-α. Acetylcholine induced a sustained increase in the immunocomplexing of calponin with HSP27 and of calponin with PKC-α in the particulate fraction, indicating an association of the translocated proteins in the membrane. To examine whether the observed interaction in vivo is due to a direct interaction of calponin with PKC-α, a cDNA of 1.3 kb of human calponin gene was PCR amplified. PCR product encoding 622 nt of calponin cDNA (nt 351–972 corresponding to amino acids 92–229) was expressed as fusion glutathione S-transferase (GST) protein in the vector pGEX -KT. We have studied the direct association of GST-calponin fusion protein with recombinant PKC-α in vitro. Western blot analysis of the fractions collected after elution with reduced glutathione buffer (pH 8.0) show a coelution of GST-calponin with PKC-α, indicating a direct association of GST-calponin with PKC-α. These data suggest that there is a direct association of translocated calponin and PKC-α in the membrane and a role for the complex calponin-PKC-α-HSP27, in contraction of colonic smooth muscle cells.

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