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1.
An. bras. dermatol ; 96(6): 712-716, Nov.-Dec. 2021. tab
Article in English | LILACS | ID: biblio-1355629

ABSTRACT

Abstract Background: The treatment of advanced periocular basal cell carcinomas becomes a challenge as surgery may involve highly mutilating procedures. Vismodegib is the first selective hedgehog inhibitor approved for the treatment of locally advanced tumors or metastatic disease. Objective: Analyze the results of treatment with vismodegib for advanced periocular basal cell carcinomas in a real-life setting of a reference center between 2014 and 2020. Methods: Retrospective longitudinal study. The patient's demographic profile, comorbidities, tumor characteristics, and treatment outcomes were analyzed. Results: A total of 13 patients were included. Median follow-up and treatment duration were 15.9 and 10.5 months, respectively. Objective clinical response rate was 76.9%: 30.8% had a complete response and 46.2% a partial response. The median duration of response was 13 months. Progressive disease was observed in 38.5% of cases, with a median of 19 months after the beginning of treatment. Eighty-four percent of the patients had at least one adverse event, and 61.54% needed to interrupt treatment temporarily or permanently to increase tolerability. Study limitations: Being a retrospective study in a real-life setting, the evaluation of objective clinical response was subjective to physician appreciation. Conclusion: Vismodegib is a safe and effective treatment for locally advanced basal cell carcinoma. To prevent recurrences, the drug should be used continually when tolerated. The role of neoadjuvant vismodegib before surgery is being investigated and might add an important step in searching for a definitive treatment for these cases.


Subject(s)
Humans , Carcinoma, Basal Cell/drug therapy , Neoplasms/drug therapy , Pyridines , Retrospective Studies , Longitudinal Studies , Hedgehog Proteins , Anilides , Neoplasm Recurrence, Local/drug therapy
2.
Article in Chinese | WPRIM | ID: wpr-880027

ABSTRACT

OBJECTIVE@#To investigate the effect of norcantharidin (NCTD) to proliferation of leukemia cells through disrupting key regulators of sonic Hedgehog (SHH) pathway and its downstream transcription factor SOX2.@*METHODS@#CCK8 was used to detected the HL60 and NB4 cells after inhibited by NCTD, SMO and GLI1 inhibitor for 24 hours. Expression level of SMO, GLI1 and SOX2 in HL60 cells with NCTD treatment was detected by immunoblot. HL60 cells were transfected with pcDNA3.1 plasmid expressing GLI1 or SOX2. Empty vector and pcDNA3. 1-EGFP were divided into negative and positive control group, respectively. The expression of exogenous GLI1 or SOX2 in HL60 cells was confirmed by immunoblot, and growth curve of HL60 cell was checked by CCK8. Proliferation of genetic modified HL60 cells treated by various dose of NCTD was detected.@*RESULTS@#NCTD, SMO/GLI1 inhibitors could inhibit the proliferation of NB4 and HL60 cells in a dose-dependent manner. Compared with solvent (DMSO)-treated control group, NCTD remarkably decreased protein level of SMO, GLI1 and SOX2. GLI1 and SOX2 were overexpressed in HL60 cells as compared with pcDNA3.1 empty vector-transfected group. Growth curve demonstrated significant proliferative advantage of GLI1/SOX2-transfected cells. CCK8 assay indicated that GLI1/SOX2-overexpressed HL60 cells were more resistant to NCTD treatment.@*CONCLUSION@#NCTD attenuates HL60 proliferation via targeting the Hedgehog/SOX2 axis.


Subject(s)
Bridged Bicyclo Compounds, Heterocyclic , Cell Proliferation , HL-60 Cells , Hedgehog Proteins , Humans , Leukemia, Myeloid, Acute , SOXB1 Transcription Factors , Zinc Finger Protein GLI1
3.
Journal of Experimental Hematology ; (6): 1504-1509, 2021.
Article in Chinese | WPRIM | ID: wpr-922286

ABSTRACT

OBJECTIVE@#To investigate the effect of arsenic disulfide (AS@*METHODS@#The human DLBCL cell OCI-LY3 was treated with different concentrations of AS@*RESULTS@#The DLBCL cell viability was decreased significantly at 24, 48 or 72 h as cultured with itraconazole. Along with the increasing of itraconazole concentration, the DLBCL cell viability was significantly reduced as compared with that in control group, and the results showed statistically significant(r=-0.690,r=-0.639, r=-0.833, r=-0.808, r=-0.578). The inhibitory and apoptosis rates of the cells were significantly increased as compared with those of the single drug-treated group after treated by the combination of itraconazole and AS@*CONCLUSION@#Itraconazole can inhibit proliferation of DLBCL cells in a concentration-and time-dependent manner. In addition, the combination of AS


Subject(s)
Apoptosis , Arsenicals , Hedgehog Proteins , Humans , Itraconazole/pharmacology , Lymphoma, Large B-Cell, Diffuse/drug therapy , Sulfides
4.
Article in Chinese | WPRIM | ID: wpr-887989

ABSTRACT

To study the mechanism of polysaccharides from seeds of Vaccaria segetalis( PSV) in the treatment of bacterial cystitis through the NLRP3 inflammasome pathway. The rat model of urinary tract infection was used and treated with PSV,and the urine and bladders were collected. The level of interleukin-10( IL-10) in rat urine was detected by enzyme linked immunosorbent assay( ELISA). Western blot and immunofluorescence staining were used to detect the expressions of sonic hedgehog( SHH) and NLRP3 inflammasome [NOD-like receptor thermoprotein domain 3( NLRP3),apoptosis associated speck like protein( ASC) and pro-caspase-1]. The expression of Toll-like receptor pathway was detected by RT-PCR. The death of 5637 cells induced by uropathogenic Escherichia coli( UPEC) and lactate dehydrogenase( LDH) release were evaluated using live/dead staining. The results showed that in the rat bladder,the expressions of SHH,NLRP3 inflammasomes and Toll-like receptors were significantly up-regulated,and NLRP3 inflammasomes were significantly activated by UPEC infection. The administration with PSV could significantly increase the concentration of IL-10 in urine,inhibit the expressions of SHH,NLRP3 inflammasomes and Toll-like receptors in bladder,and inhibit the activation of NLRP3 inflammasomes. A large number of 5637 cells were dead after UPEC infection and caused LDH production. PSV could significantly inhibit the death of 5637 cells and the release of LDH. In conclusion,PSV could inhibit the expression and activation of NLRP3 inflammasomes by inhibiting the Toll-like receptor pathway,thereby mitigating the bladder injury.


Subject(s)
Animals , Hedgehog Proteins , Inflammasomes/genetics , Interleukin-1beta , NLR Family, Pyrin Domain-Containing 3 Protein/genetics , Polysaccharides/pharmacology , Rats , Seeds , Urinary Bladder , Urinary Tract Infections/drug therapy , Vaccaria
5.
Article in English | WPRIM | ID: wpr-880634

ABSTRACT

OBJECTIVES@#Silence of SET domain containing lysine methyltransferase 7 (SET7) alleviates myocardial tissue injury caused by ischemia-reperfusion. But the effects of SET7 on angiotensin II (Ang II)-induced myocardial fibroblast proliferation and the collagen synthesis are not clear. The purpose of this study was to explore the effect of SET7 on the proliferation and collagen synthesis of myocardial fibroblasts and its mechanisms.@*METHODS@#Myocardial fibroblasts were isolated and identified by immunofluorescence. Myocardial fibroblasts were randomly divided into 4 groups: a control group (cells were normally cultured), an Ang II group (cells were treated with 100 nmol/L Ang II for 24 h), a siCtrl group (cells were transfected with siRNA control and were then treated with 100 nmol/L Ang II for 24 h), and a siSET7 group (cells were transfected with siRNA SET7 and were then treated with 100 nmol/L Ang II for 24 h). Cell counting kit-8 (CCK-8) and 5-ethynyl-2'-deoxyuridine (EdU) assay were used to evaluate cell proliferation. Real-time PCR was used to detect the mRNA levels of SET7, collagen I, collagen III, and α-smooth muscle actin (α-SMA). Western blotting was used to detect the protein expression of SET7, collagen I, collagen III, α-SMA, sonic hedgehog (Shh), ptched1 (Ptch1), and glioma-associated oncogene homolog 1 (Gli1).@*RESULTS@#Fluorescence microscopy showed positive vimentin staining, and myocardial fibroblasts were in good condition. As compared to the control group, the mRNA and protein levels of SET7 in the Ang II group were significantly upregulated; cell proliferation rate and EdU fluorescence intensity in the Ang II group were significantly increased; the mRNA and protein levels of collagen I, collagen III, and α-SMA were significantly upregulated (all @*CONCLUSIONS@#Silence of SET7 gene inhibits Ang II-induced proliferation and collagen synthesis of myocardial fibroblasts. Shh signaling pathway may be involved in this process.


Subject(s)
Angiotensin II/pharmacology , Cell Proliferation , Cells, Cultured , Collagen/genetics , Fibroblasts , Hedgehog Proteins
6.
Article in Chinese | WPRIM | ID: wpr-879025

ABSTRACT

To prove that ursolic acid(UA)could activate the autophagy of colorectal cancer HCT116 cells by inhibiting hedgehog signaling pathway. The effect of UA on the viability of HCT116 cells was determined by MTT assay. The effect of UA on the proliferation and migration of HCT116 cells was detected by crystal violet staining and scratch test. In the study on autophagy, the time points were screened out first: the autophagy fluorescence intensity of UA acting on HCT116 at different time points were detected by Cell Meter~(TM) Autophagy Assay Kit; Western blot was used to detect the expression of autophagy protein P62 at different time points. Then, Cell Meter~(TM) Autophagy Assay Kit was used to detect the effect of UA on autophagy fluorescence intensity of HCT116 cells. The effect of different doses of UA on the expressions of LC3Ⅱ and P62 proteins in HCT116 cells were detected by Western blot. Further, AdPlus-mCherry-GFP-LC3 B adenovirus transfection was used to detect the effects of UA on autophagy flux of HCT116 cells; UA combined with autophagy inhibitor chloroquine(CQ) was used to detect the expression of LC3Ⅱ by Western blot. In terms of mechanism, the effect of UA on hedgehog signaling pathway-related proteins in HCT116 cells was detected by Western blot. The results showed that UA inhibited the activity, proliferation and migration of HCT116 cells. UA enhanced the fluorescence intensity of autophagy in HCT116 cells, while promoting the expression of LC3Ⅱ and inhibiting the expression of P62, in a time and dose dependent manner. UA activated the autophagy in HCT116 cells, which manifested that UA resulted in the accumulation of fluorescence spots and strengthened the fluorescence intensity of autophagosomes; compared with UA alone, UA combined with autophagy inhibitor CQ promoted the expression of LC3Ⅱ. UA reduced the expressions of PTCH1, GLI1, SMO, SHH and c-Myc in hedgehog signaling pathway, while increased the expression of Sufu. In conclusion, our study showed that UA activated autophagy in colorectal cancer HCT116 cells, which was related to the mechanism in inhibiting hedgehog signaling pathway activity.


Subject(s)
Apoptosis , Autophagy , Cell Line, Tumor , Colorectal Neoplasms , Hedgehog Proteins/genetics , Humans , Signal Transduction , Triterpenes
7.
Electron. j. biotechnol ; 46: 30-37, jul. 2020. tab, graf
Article in English | LILACS | ID: biblio-1223233

ABSTRACT

BACKGROUND: The effects of dietary nutrition on tail fat deposition and the correlation between production performance and the Hh signaling pathway and OXCT1 were investigated in fat-tailed sheep. Tan sheep were fed different nutritional diets and the variances in tail length, width, thickness and tail weight as well as the mRNA expression of fat-related genes (C/EBPα, FAS, LPL, and HSL) were determined in the tail fat of sheep at three different growth stages based on their body weight. Furthermore, the correlations between tail phenotypes and the Hedgehog (Hh) signaling pathway components (IHH, PTCH1, SMO, and GLI1) and OXCT1 were investigated. RESULTS: C/EBPα, FAS, LPL, and HSL were expressed with differences in tail fat of sheep fed different nutritional diets at three different growth stages. The results of the two-way ANOVA showed the significant effect of nutrition, stage, and interaction on gene expression, except the between C/EBPα and growth stage. C/EBPα, FAS, and LPL were considerably correlated with the tail phenotypes. Furthermore, the results of the correlation analysis demonstrated a close relationship between the tail phenotypes and Hh signaling pathway and OXCT1. CONCLUSIONS: The present study demonstrated the gene-level role of dietary nutrition in promoting tail fat deposition and related tail fat-related genes. It provides a molecular basis by which nutritional balance and tail fat formation can be investigated and additional genes can be identified. The findings of the present study may help improve the production efficiency of fat-tailed sheep and identify crucial genes associated with tail fat deposition.


Subject(s)
Animals , Tail/metabolism , Sheep/genetics , Adipose Tissue , Diet , Phenotype , RNA, Messenger , Coenzyme A-Transferases , Gene Expression , Body Fat Distribution , Adipogenesis , Lipogenesis/genetics , Hedgehog Proteins/genetics , Real-Time Polymerase Chain Reaction
8.
Journal of Experimental Hematology ; (6): 1298-1302, 2020.
Article in Chinese | WPRIM | ID: wpr-827123

ABSTRACT

OBJECTIVE@#To study the effect of SMO inhibitor (Jervine) on proliferation, apoptosis and cell cycle of MDS cell line MUTZ-1, and its mechanism.@*METHODS@#The effect of different concentrations Jervine on proliferation of MUTZ-1 cells was detected by CCK-8 method. Apoptosis and cell cycle of MUTZ-1 cells were detected by flow cytometry. Western blot was used to detect the changes of Shh signaling pathway effecting proteins BCL2 and CyclinD1. The expression levels of Smo and Gli1 gene were detected by real-time fluorescent quantitative polymerase chain reaction (RT-qPCR).@*RESULTS@#Jervine inhibited MUTZ-1 cell proliferation in a concentration dependent manner (24 h, r=-0.977), the apoptosis rate of MUTZ-1 cells increased with the enhancement of concentration of Jervine in MUTZ-1 cells (P<0.001), the cell proportion of G phase increased and the cell number of S phase decreased with enhancement of concentration (P<0.001). The result of RT-qPCR and Western blot showed that the expression of Smo, Gli1 mRNA and BCL2, CyclinD1 proteins decreased (P<0.05).@*CONCLUSION@#SMO inhibitor can effectively inhibit the growth of MDS cell line MUTZ-1 improve the cell apoptosis and induce cell cycle arrest. Its action mechanism may be related with dowm-regulating the expression of BCL2 and CyclinD1.


Subject(s)
Apoptosis , Cell Line, Tumor , Cell Proliferation , Hedgehog Proteins , Humans , Myelodysplastic Syndromes , Signal Transduction , Veratrum Alkaloids
10.
Article in English | WPRIM | ID: wpr-772278

ABSTRACT

Odontogenic keratocysts (OKCs) are common cystic lesions of odontogenic epithelial origin that can occur sporadically or in association with naevoid basal cell carcinoma syndrome (NBCCS). OKCs are locally aggressive, cause marked destruction of the jaw bones and have a propensity to recur. PTCH1 mutations (at ∼80%) are frequently detected in the epithelia of both NBCCS-related and sporadic OKCs, suggesting that PTCH1 inactivation might constitutively activate sonic hedgehog (SHH) signalling and play a major role in disease pathogenesis. Thus, small molecule inhibitors of SHH signalling might represent a new treatment strategy for OKCs. However, studies on the molecular mechanisms associated with OKCs have been hampered by limited epithelial cell yields during OKC explant culture. Here, we constructed an isogenic PTCH1 cellular model of PTCH1 inactivation by introducing a heterozygous mutation, namely, c.403C>T (p.R135X), which has been identified in OKC patients, into a human embryonic stem cell line using the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) system. This was followed by the induction of epithelial differentiation. Using this in vitro isogenic cellular model, we verified that the PTCH1 heterozygous mutation causes ligand-independent activation of SHH signalling due to PTCH1 haploinsufficiency. This activation was found to be downregulated in a dose-dependent manner by the SHH pathway inhibitor GDC-0449. In addition, through inhibition of activated SHH signalling, the enhanced proliferation observed in these induced cells was suppressed, suggesting that GDC-0449 might represent an effective inhibitor of the SHH pathway for use during OKC treatment.


Subject(s)
Anilides , Pharmacology , Basal Cell Nevus Syndrome , Hedgehog Proteins , Genetics , Pharmacology , Humans , Molecular Targeted Therapy , Odontogenic Cysts , Genetics , Therapeutics , Odontogenic Tumors , Genetics , Therapeutics , Pyridines , Pharmacology
11.
Article in English | WPRIM | ID: wpr-772274

ABSTRACT

The Hedgehog (Hh) signalling pathway is essential for cellular proliferation and differentiation during embryonic development. Gain and loss of function of Hh signalling are known to result in an array of craniofacial malformations. To determine the critical period for Hh pathway antagonist-induced frontal bone hypoplasia, we examined patterns of dysmorphology caused by Hh signalling inhibition. Pregnant mice received a single oral administration of Hh signalling inhibitor GDC-0449 at 100 mg•kg or 150 mg•kg body weight at preselected time points between embryonic days (E)8.5 and 12.5. The optimal teratogenic concentration of GDC-0449 was determined to be 150 mg•kg. Exposure between E9.5 and E10.5 induced frontal bone dysplasia, micrognathia and limb defects, with administration at E10.5 producing the most pronounced effects. This model showed decreased ossification of the frontal bone with downregulation of Hh signalling. The osteoid thickness of the frontal bone was significantly reduced. The amount of neural crest-derived frontal bone primordium was reduced after GDC-0449 exposure owing to a decreased rate of cell proliferation and increased cell death.


Subject(s)
Administration, Oral , Anilides , Pharmacology , Animals , Bone Diseases, Developmental , Cell Proliferation , Physiology , Female , Frontal Bone , Congenital Abnormalities , Hedgehog Proteins , Limb Deformities, Congenital , Mice , Micrognathism , Osteogenesis , Pregnancy , Pyridines , Pharmacology , Signal Transduction
12.
Journal of Experimental Hematology ; (6): 1402-1408, 2019.
Article in Chinese | WPRIM | ID: wpr-775707

ABSTRACT

OBJECTIVE@#To investigate the mechanism of rapamycin-induced apoptosis of chronic myelogenous leukemia cells.@*METHODS@#The chronic granulocytic leukemia K562 cells were divided into 3 groups: A, B and C group were treated with rapamycin of 10, 15 and 20 nmol/L, repectively for 24 h, while the K562 cells in control group were not treated with rapamycin. The effect of rapamycin on the proliferation of K562 cells was detected by MTT, and the effect of rapamycin on the apoptosis of K562 cells was detected by AnnexinV-FITC/PI double staining. The expression level of EZH2/Hedgehog signaling pathway genes in K562 cells was detected by RT-PCR, and Western blot was used to detect the levels of apoptotic protein and the related signaling pathway proteins in K562 cells.@*RESULTS@#The MTT assay showed that the different concentration of rapamycin had obvious inhibitory effects on the cells, and the survival rate of cells in group C was 37.6%±3.4%, which was significantly lower than that of the other groups (P<0.05). The apoptosis rate of cells in group C was 93.1%±8.1%, which was significantly higher than that of the other groups (P<0.05). By Western blot, it was found that the relative expression levels of Caspase-3 and BAX protein in group C were 0.36 ± 0.04 and 0.39±0.06, respectively, which were significantly higher than those in other groups (P<0.05), and the level of BCL-2 protein was 0.17±0.03, which was significantly lower than that of other groups (P<0.05). By RT-PCR, it was found that the mRNA levels of EZH2 and Hedgehog genes in A, B and C groups were significantly lower than those in the control group (P<0.05), but mRNA level of Ptch1 gene was significantly higher than that of the control (P<0.05). By Western blot, it was found that the expression levels of EZH2 and Hedgehog protein in A, B and C groups were significantly lower than that in the control group (P<0.05), but the level of Ptch1 protein was higher than that of the control (P<0.05). The relative levels of EZH2 and Hedgehog protein in group C were 0.21 ±0.03 and 0.16±0.05 respectively, which were significantly lower than those in other groups (P<0.05), and Ptch1 protein level were 0.46 ±0.06, significantly higher than that of other groups (P<0.05).@*CONCLUSION@#Rapamycin can inhibit the protein expression of EZH2 in leukemic cells, thus interfere with the activation of Hedgehog signaling pathway, promote the expression of apoptotic protein, reduce the level of anti apoptotic protein, and eventually induce apoptosis of leukemia cells.


Subject(s)
Apoptosis , Cell Proliferation , Enhancer of Zeste Homolog 2 Protein , Hedgehog Proteins , Humans , K562 Cells , Signal Transduction , Sirolimus
13.
Article in Chinese | WPRIM | ID: wpr-774047

ABSTRACT

OBJECTIVE@#To study the expression of Shh and Wnt5a genes in the limb buds of NIPBL fetal rats and the association of these two genes with Cornelia de Lange syndrome (CdLS).@*METHODS@#A total of 72 NIPBL fetal rats were divided into an experimental group and a control group, with 36 rats in each group. The limb buds were collected from 12 fetal rats each on embryonic days 10, 11 and 12 (E10, E11 and E12) respectively. Real-time PCR and Western blot were used to measure the mRNA and protein expression of Shh and Wnt5a.@*RESULTS@#The mRNA and protein expression of Shh and Wnt5a was detected in the limb buds on E10, E11 and E12, and the experimental group had significantly lower expression than the control group (P<0.01). The mRNA and protein expression of Shh and Wnt5a in limb buds was at a low level on E10, followed by an increase on E11 and a reduction on E12, and the expression on E12 was still lower than that on E10 (P<0.01).@*CONCLUSIONS@#The mRNA and protein expression of Shh and Wnt5a are consistent. The pathogenesis of CdLS may be associated with the low mRNA and protein expression of Shh and Wnt5a inhibited by the low expression of NIPBL gene.


Subject(s)
Animals , De Lange Syndrome , Hedgehog Proteins , Mutation , Phenotype , Proteins , RNA, Messenger , Rats , Wnt-5a Protein
14.
Yonsei Medical Journal ; : 898-904, 2019.
Article in English | WPRIM | ID: wpr-762043

ABSTRACT

PURPOSE: Sonic hedgehog (Shh) signaling pathway is known to play a crucial role in carcinogenesis in various malignancies, including lung cancer regarding tumorigenesis, angiogenesis, and cellular differentiation. The aim of this study was to investigate the value of components of Shh pathway as a prognostic marker in extensive stage small cell lung cancer (ES-SCLC) patients. MATERIALS AND METHODS: We retrospectively analyzed data of 36 patients who were diagnosed with ES-SCLC between 2008 and 2012 at a single center. We performed immuo-histochemistry for glioma-associated oncogene homolog zinc finger protein 1 (Gli1), patched, Shh, and Ptch-mediated repression of smoothened (Smo) proteins using formalin-fixed, paraffin-embedded tissue derived from primary tumors. We then conducted survival analysis to evaluate the prognostic impact of these markers. RESULTS: All 36 patients received platinum-based doublet chemotherapy. The median progression free survival and median overall survival were 6.9 months [95% confidence interval (CI), 6.5–7.3] and 11.7 months (95% CI, 9.1–14.3), respectively. The overall response rate was 84%. Of the 36 tissue specimens examined, over-expression of Gli1, Patched, Shh, and Smo was found in 12 (33.3%), five (13.9%), five (13.9%), and six (16.7%) cases, respectively. We found that high expression of Shh was associated with worse progression free survival (6.3 vs. 7.6 months, p=0.005) and overall survival (9.2 vs. 12.0 months, p=0.039) by both univariate and multivariate analyses, whereas other markers were not related to patient prognosis. CONCLUSION: A high proportion of small cell lung cancer tumors express proteins related to Shh pathway, and over-expression of Shh is correlated with poor prognosis.


Subject(s)
Carcinogenesis , Disease-Free Survival , Drug Therapy , Hedgehog Proteins , Hedgehogs , Humans , Lung Neoplasms , Multivariate Analysis , Oncogenes , Prognosis , Repression, Psychology , Retrospective Studies , Small Cell Lung Carcinoma , Zinc Fingers
15.
Int. j. morphol ; 37(1): 221-226, 2019. tab, graf
Article in Spanish | LILACS | ID: biblio-990030

ABSTRACT

RESUMEN: Para que se desarrolle el iris, se requiere una especificación de la capa periférica de la copa óptica a un destino no neuronal y además la migración de células mesenquimales perioculares. Nuestro objetivo fue reconocer los cambios histológicos de los derivados periféricos de la copa óptica y mesénquima periocular, como también reconocer la presencia del morfógeno Sonic hedgehog (Shh) en las capas que constituyen el esbozo de iris. Se utilizaron 15 ratones hembras (Mus musculus) adultas jóvenes gestantes. Se realizó eutanasia con tiopental sódico. Los embriones y fetos de 12, 14,5 y 17 días post-coital (dpc) fueron procesados con técnica histológica e inmunohistoquímica con anticuerpo anti-Shh (scbt, H-160, conejo) con dilución 1:100 en PBS. A los 12 dpc, se observa una cópa óptica que presenta capas retinianas interna y externa, y el iris no se observa. Entre el cristalino y el ectodermo superficial se identifican 4 capas de células mesenquimales. A los 14,5 dpc, el iris contiene dos capas epiteliales (interna y externa) que se continúan con las capas neural y pigmentaria de la retina. Se observan 8 capas de células mesenquimales. A los 17 dpc, la capa epitelial interna del iris presenta un segmento más elongado con inmunotinción positiva a Shh y otra parte que constituye un epitelio de células cilíndricas simples negativas a este anticuerpo. La capa epitelial externa presenta el mismo epitelio inmunonegativo. Las capas de la retina también son positivas, como también la periferia del cristalino. No esta formado el iris ni tampoco el cuerpo ciliar. La inmunopositividad en el cristalino, en el primer segmento de la capa interna del esbozo del iris y en la capa ganglionar retinal a los 17 dpc, se relaciona con la diferenciación tardía del iris y con los ojos cerrados de las crías al nacimiento.


SUMMARY: In order for the iris to develop, a specification of the peripheral layer of the optic cup to a non-neuronal target is required, as well as the migration of periocular mesenchymal cells. Our aim was to recognize the histological changes of peripheral derivatives of the optic cup and periocular mesenchyme, as well as recognize the presence of the morphogen Sonic hedgehog (Shh) in the layers constituting the outline of the iris. 15 female mice (Mus musculus) pregnant young adults were used. Euthanasia was performed with sodium thiopental. Embryos and fetuses of 12, 14.5 and 17 days post-coital (dpc) were processed with histological and immunohistochemical technique with anti-Shh antibody (scbt, H 160, rabbit) with dilution 1:100 in PBS. At 12 dpc, an optic cup showing internal and external retinal layers is observed, and the iris is not observed. Between the lens and the superficial ectoderm, 4 layers of mesenchymal cells are identified. At 14.5 dpc, the iris contains two epithelial layers (internal and external) that are continued with the neural and pigmentary layers of the retina. 8 layers of mesenchymal cells are observed. At 17 dpc, the inner epithelial layer of the iris presents a more elongated segment with positive immunostaining to Shh and another part that constitutes an epithelium of simple cylindrical cells negative to this antibody. The outer epithelial layer presents the same immunonegative epithelium. The layers of the retina are also positive, as well as the periphery of the lens. The iris is not formed nor is the ciliary body.The immunopositivity in the lens, in the first segment of the inner layer of the iris outline and in the retinal ganglion layer at 17 dpc, is related to the late differentiation of the iris and the closed eyes of the offspring at birth.


Subject(s)
Animals , Female , Mice , Iris/embryology , Eye/embryology , Hedgehog Proteins , Iris/anatomy & histology , Eye/anatomy & histology , Morphogenesis
16.
Int. j. morphol ; 36(2): 500-506, jun. 2018. tab, graf
Article in Spanish | LILACS | ID: biblio-954144

ABSTRACT

Cada término incluido en Terminologia Embryologica (TE), publicada en 2013 y con una segunda edicición en 2017 sujeta a aprobación por la Federación Internacional de Programas de Terminología Anatómica (FIPAT), se debe estructurar en base a sus principios: los nombres deben tener un valor informativo, se suprimen los epónimos, homónimos y nombres concordantes. Sin embargo, esto no ocurre en algunos términos. Es por ello que se analizaron los términos incluidos en el ítem "Factores de Crecimiento" (Factores crescentiae, E.4.0.1.0.0.0.1) en la forma en que se presentan, se contrastó su concordancia con lo publicado en revisiones de la base de datos PubMed y se describió la forma de nomenclatura de ellos en base a lo utilizado por el Comité de Nomenclatura de Genes de la Organización del Genoma Humano (HGNC). Se evidenció que en los términos Familia Hedgehog (Familia erinacea, E.4.0.1.0.0.0.6) y Superfamilia del Factor de Crecimiento Epidermal (EGF) (Superfamilia factoris epidermalis [EGF] crescentiae, E.4.0.1.0.0.0.10) no concuerdan con la clasificación propuesta por TE en base al receptor que se ve involucrado en la actividad del factor de crecimiento, ya que en el caso de la familia Hedgehog no sólo participa el receptor patched, sino también el smoothened. En el EGF hay actividad del receptor tirosina kinasa y no del serina/treonina kinasa, como se presenta en el documento oficial. También, aparecen los ligandos fibronectina/laminina y delta/serrate como factores de crecimiento, pese a no ser catalogados como tal. Por otra parte, la forma en la que se construyen este tipo de términos muchas veces no es como la plantea la FIPAT, sino que obedece a la línea de trabajo que del HGNC. TE debiese modificar el criterio empleado en el ítem factores de crecimiento.


Each term included in Terminologia Embryologica (TE), published in 2013 and with a second edition in 2017 subject to approval by the Federative International Programme For Anatomical Terminology (FIPAT), must be structured based on its principles: Names must have an informative value, eponyms, homonyms and concordant names are deleted. However, this does not happen in some terms. That is why the terms included in the section "Growth Factors" (Factores crescentiae, E.4.0.1.0.0.0.1) were analyzed in the way they are presented, their concordance with what was published in reviews of PubMed database, and their naming form was described based on what was used by the Genes Nomenclature Committee of the Human Genome Organisation (HGNC). It was evidenced that in the terms Family Hedgehog (Familia erinacea, E.4.0.1.0.0.0.6) and Superfamily of the Epidermal Growth Factor (EGF) (Superfamilia factoris epidermalis [EGF] crescentiae, E.4.0.1.0.0.0.10) these do not agree with the classification proposed by TE based on the receiver that is involved in the activity of the growth factor, since in the case of Hedgehog family not only does the patched receptor participate, but the smoothened one does as well. In EGF there is tyrosine kinase receptor activity and not serine/threonine kinase, as presented in the official document. Also, the ligands fibronectin/laminin and delta/serrate appear as growth factors, despite not being catalogued as such. Furthermore, oftentimes the way in which such terms are constructed is contrary to the FIPAT presentation, but follows the HGNC line of work that holds the HGNC.


Subject(s)
Embryology , Intercellular Signaling Peptides and Proteins , Terminology as Topic , Epidermal Growth Factor , Hedgehog Proteins
17.
Int. j. morphol ; 36(2): 693-698, jun. 2018. graf
Article in Spanish | LILACS | ID: biblio-954173

ABSTRACT

Sonic hedgehog (Shh) es un morfógeno esencial para el desarrollo de diversas estructuras, tales como notocorda, placa del piso del tubo neural, miembros, entre otros. Se buscó determinar la inmunolocalización de Shh en embriones y fetos de ratón. Para ello, se eutanasiaron 10 ratones gestantes (Mus musculus) BALB/c, un grupo de 5 animales a los 12,5 días post-coito (dpc), y otro grupo a los 17,5 dpc. Los embriones y fetos obtenidos fueron fijados en formalina al 10 % tamponada en PBS e incluidos en paraplast. Se realizaron cortes transversales seriados. Se utilizó anticuerpo policlonal Shh (Santa Cruz Biotechnology, H-160, conejo), dilución 1:100. Se identificó y describió la inmunolocalización de las muestras marcadas positivamente. La expresión de Shh en los embriones de 12,5 dpc fue inmunopositiva en notocorda, placa del piso del tubo neural, precartílago de radio y ulna, y prácticamente todos los epitelios: bronquial, intestinal, vejiga y uretra. En la etapa fetal, a los 17,5 dpc la inmunopositividad desaparece en el cartílago a excepción de zonas de osificación, disminuye en la epidermis pero aparece en folículos pilosos. La mucosa intestinal se ha diferenciado en segmentos, mostrando una inmunotinción mayor a nivel de las vellosidades intestinales. Shh actúa en distintos estadios del periodo gestacional, siendo clave en la diferenciación de distintas estructuras. En etapas embrionaria, es vital en la formación del sistema nervioso, organogénesis y formación de miembros, por lo que su expresión se encuentra en estas zonas. Sin embargo, en la etapa fetal la expresión cambia a estructuras de mayor especialización como folículo piloso y vellosidades intestinales.


Sonic hedgehog (Shh) is an essential morphogen for the development of various structures, such as notochord, neural tube floor plate, limbs, among others. We sought to determine the immunolocalization of Shh in embryos and mouse fetuses. To do this, 10 pregnant mice (Mus musculus) BALB /c were euthanized, a group of 5 animals at 12.5 days postcoitus (dpc), and another group at 17.5 dpc. Embryos and fetuses obtained were fixed in 10 % formalin buffered in PBS and embedded in paraplast. Serial cross sections were made. Polyclonal antibody Shh (Santa Cruz Biotechnology, H-160, rabbit), dilution 1:100 was used. The immunolocalization of the positively labeled samples was identified and described. Shh expression in 12.5 dpc embryos was immunopositive in notochord, neural tube floor plate, radius precartilage and ulna, and practically all epithelia: bronchial, intestinal, bladder and urethra. In the fetal stage, at 17.5 dpc the immunopositivity disappears in the cartilage except for areas of ossification, decreases in the epidermis but appears in hair follicles. The intestinal mucosa has differentiated into segments, showing greater immunostaining at the level of the intestinal villi. Shh acts in different stages of the gestational period, being key in the differentiation of different structures. In embryonic stages, it is vital in the formation of the nervous system, organogenesis and formation of limbs, so its expression is found in these areas. However, in the fetal stage the expression changes to more specialized structures such as hair follicles and intestinal villi.


Subject(s)
Animals , Female , Mice , Organogenesis/physiology , Hedgehog Proteins/metabolism , Embryonic and Fetal Development , Immunohistochemistry , Embryo, Mammalian , Mice, Inbred BALB C
18.
Chinese Medical Journal ; (24): 1191-1198, 2018.
Article in English | WPRIM | ID: wpr-688146

ABSTRACT

<p><b>Background</b>The hedgehog signaling system (HHS) plays an important role in the regulation of cell proliferation and differentiation during the embryonic phases. However, little is known about the involvement of HHS in the malignant transformation of cells. This study aimed to detect the role of HHS in the malignant transformation of human bronchial epithelial (16HBE) cells.</p><p><b>Methods</b>In this study, two microfluidic chips were designed to investigate cigarette smoke extract (CSE)-induced malignant transformation of cells. Chip A contained a concentration gradient generator, while chip B had four cell chambers with a central channel. The 16HBE cells cultured in chip A were used to determine the optimal concentration of CSE for inducing malignant transformation. The 16HBE cells in chip B were cultured with 12.25% CSE (Group A), 12.25% CSE + 5 μmol/L cyclopamine (Group B), or normal complete medium as control for 8 months (Group C), to establish the in vitro lung inflammatory-cancer transformation model. The transformed cells were inoculated into 20 nude mice as cells alone (Group 1) or cells with cyclopamine (Group 2) for tumorigenesis testing. Expression of HHS proteins was detected by Western blot. Data were expressed as mean ± standard deviation. The t-test was used for paired samples, and the difference among groups was analyzed using a one-way analysis of variance.</p><p><b>Results</b>The optimal concentration of CSE was 12.25%. Expression of HHS proteins increased during the process of malignant transformation (Group B vs. Group A, F = 7.65, P < 0.05). After CSE exposure for 8 months, there were significant changes in cellular morphology, which allowed the transformed cells to grow into tumors in 40 days after being inoculated into nude mice. Cyclopamine could effectively depress the expression of HHS proteins (Group C vs. Group B, F = 6.47, P < 0.05) and prevent tumor growth in nude mice (Group 2 vs. Group 1, t = 31.59, P < 0.01).</p><p><b>Conclusions</b>The activity of HHS is upregulated during the CSE-induced malignant transformation of 16HBE cells. Cyclopamine can effectively depress expression of HHS proteins in vitro and prevent tumor growth of the transformed cells in vivo.</p>


Subject(s)
Animals , Cell Transformation, Neoplastic , Genetics , Metabolism , Gene Expression Regulation, Neoplastic , Genetics , Physiology , Hedgehog Proteins , Genetics , Metabolism , Lab-On-A-Chip Devices , Mice , Mice, Inbred BALB C , Mice, Nude , Microfluidics , Signal Transduction , Genetics , Physiology , Smoke , Smoking
19.
An. bras. dermatol ; 92(4): 517-520, July-Aug. 2017.
Article in English | LILACS | ID: biblio-886982

ABSTRACT

Abstract: Basal cell carcinoma is the most common cancer, presenting low mortality but high morbidity, and it has as risk factor exposure to sunlight, especially UVB spectrum. The most important constitutional risk factors for basal cell carcinoma development are clear phototypes (I and II, Fitzpatrick classification), family history of basal cell carcinoma (30-60%), freckles in childhood, eyes and light hair. The environmental risk factor better established is exposure to ultraviolet radiation. However, different solar exposure scenarios probably are independent risk factors for certain clinical and histological types, topographies and prognosis of this tumor, and focus of controversy among researchers. Studies confirm that changes in cellular genes Hedgehog signaling pathway are associated with the development of basal cell carcinoma. The cellular Hedgehog signaling pathway is activated in organogenesis, but is altered in various types of tumors.


Subject(s)
Humans , Skin Neoplasms/genetics , Carcinoma, Basal Cell/genetics , Hedgehog Proteins/physiology , Hedgehog Proteins/genetics
20.
Article in Chinese | WPRIM | ID: wpr-357508

ABSTRACT

<p><b>OBJECTIVE</b>To determine the influence of altered masticatory loading on Indian hedgehog (Ihh)-parathyroid hormone-like related protein (PThrP) pathway in the condylar cartilage of growing rabbits.</p><p><b>METHODS</b>A total of 48 10-day-old rabbits were randomly divided into two groups and fed different kinds of food, such as solid diet and soft diet. The animals were sacrificed after 2, 4, 6, and 8 weeks. Difference of Ihh and PThrP expression levels induced by altered masticatory loading was tested by hematoxylin-eosin (HE), immunohistochemistry, Western blot, and real-time polymerase chain reaction (PCR).</p><p><b>RESULTS</b>The thickness of condylar cartilage and expression levels of Ihh and PThrP proteins and mRNA of the solid diet groups exceeded those of the soft diet groups. The decreasing tendencies of the expression levels of Ihh and PThrP proteins and mRNA were observed at 2, 4, 6, 8 weeks.</p><p><b>CONCLUSIONS</b>Low masticatory loading may delay or inhibit the development of condylar cartilage and its growing factors Ihh and PThrP. Therefore, masticatory loading plays an important role in the development of condylar cartilage.</p>


Subject(s)
Animals , Blotting, Western , Cartilage , Chondrocytes , Hedgehog Proteins , Immunohistochemistry , Mastication , Parathyroid Hormone , Rabbits , Real-Time Polymerase Chain Reaction
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