Establishment and characterization of bone marrow mesenchymal stem cell lines stably synthesizing high-level dopamine / 生物工程学报

A triple-transgenic (tyrosine hydroxylase/dopamine decarboxylase/GTP cyclohydrolase 1, TH/DDC/GCH1) bone marrow mesenchymal stem cell line (BMSCs) capable of stably synthesizing dopamine (DA) transmitters were established to provide experimental evidence for the clinical treatment of Parkinson's disease (PD) by using this cell line. The DA-BMSCs cell line that could stably synthesize and secrete DA transmitters was established by using the triple transgenic recombinant lentivirus. The triple transgenes (TH/DDC/GCH1) expression in DA-BMSCs was detected using reverse transcription-polymerase chain reaction (RT-PCR), Western blotting, and immunofluorescence. Moreover, the secretion of DA was tested by enzyme-linked immunosorbent assay (ELISA) and high-performance liquid chromatography (HPLC). Chromosome G-banding analysis was used to detect the genetic stability of DA-BMSCs. Subsequently, the DA-BMSCs were stereotactically transplanted into the right medial forebrain bundle (MFB) of Parkinson's rat models to detect their survival and differentiation in the intracerebral microenvironment of PD rats. Apomorphine (APO)-induced rotation test was used to detect the improvement of motor dysfunction in PD rat models with cell transplantation. The TH, DDC and GCH1 were expressed stably and efficiently in the DA-BMSCs cell line, but not expressed in the normal rat BMSCs. The concentration of DA in the cell culture supernatant of the triple transgenic group (DA-BMSCs) and the LV-TH group was extremely significantly higher than that of the standard BMSCs control group (P < 0.000 1). After passage, DA-BMSCs stably produced DA. Karyotype G-banding analysis showed that the vast majority of DA-BMSCs maintained normal diploid karyotypes (94.5%). Moreover, after 4 weeks of transplantation into the brain of PD rats, DA-BMSCs significantly improved the movement disorder of PD rat models, survived in a large amount in the brain microenvironment, differentiated into TH-positive and GFAP-positive cells, and upregulated the DA level in the injured area of the brain. The triple-transgenic DA-BMSCs cell line that stably produced DA, survived in large numbers, and differentiated in the rat brain was successfully established, laying a foundation for the treatment of PD using engineered culture and transplantation of DA-BMSCs.

Caracterización clínica de pacientes chilenos con displasia fibrosa/síndrome de McCune-Albright / Clinical features of Chilean patients with Fibrous Dysplasia/McCune-Albright Syndrome

BACKGROUND: Fibrous Dysplasia/McCune-Albright Syndrome (FD/MAS) is characterized by a spectrum of manifestations that may include fibrous dysplasia of bone and multiple endocrinopathies. AIM: To describe the clinical spectrum, the study and follow-up of patients with FD/MAS cared at our institution. MATERIAL AND METHODS: Review of medical records of 12 pediatric and adult patients (11 women) who met the clinical and genetic diagnostic criteria for FD/ MAS. RESULTS: The patients' mean age at diagnosis was 4.9 ± 5.5 years. The most common initial clinical manifestation was peripheral precocious puberty (PPP) in 67% of patients and 75% had café-au-lait spots. Fibrous dysplasia was present in 75% of patients and the mean age at diagnosis was 7.9 ± 4.7 years. Ten patients had a bone scintigraphy, with an age at the first examination that varied between 2 and 38 years of age. The most frequent location of dysplasia was craniofacial and appendicular. No patient had a recorded history of cholestasis, hepatitis, or pancreatitis. In four patients, a genetic study was performed that was positive for the pathogenic variant of guanine nucleotide binding protein, alpha stimulating (GNAS). CONCLUSIONS: These patients demonstrate the variable nature of the clinical presentation and study of FD/MAS. It is essential to increase the index of diagnostic suspicion and adherence to international recommendations.

Distonías primarias respondedoras a levodopa (DRD): búsqueda sistemática en Latinoamérica / Dopa-responsive dystonia (DRD): systematic search in Latin America

RESUMEN Las distonías que responden a levodopa (DRD, siglas en inglés) abarcan un grupo de distonías primarias, causadas por deficiencias enzimáticas en la vía metabólica de las aminas y, por definición, comparten como característica principal su respuesta favorable y sostenida a levodopa. Existen hasta seis genes asociados a DRD, siendo el gen GCH1 el más frecuentemente involucrado. La presentación típica de esta entidad se caracteriza por su aparición en la niñez, distonía de inicio en miembros inferiores con fluctuación diurna, leve parkinsonismo y respuesta clara a dosis bajas de levodopa. Se incluye una búsqueda sistemática de la literatura con casos de DRD publicados en Latinoamérica.

SUMMARY Dopa-responsive dystonia (DRD) encompasses a heterogenous group of primary dystonias, caused by enzymatic deficiencies across the amines pathway and, by definition, show as their main characteristic a favorable and sustained response to levodopa. There are up to 6 genes associated with DRD, including pathogenic variants of the GCH1 gene as the most frequently involved. The typical presentation of DRD is characterized by start in childhood, lower limb-onset dystonia with daytime fluctuation, mild parkinsonism, and a sustained response to low doses of levodopa. A systematic literature search on DRD reported cases in Latin America is presented.

Era-like GTP protein gene expression in rice / Expressão do gene da proteína GTP semelhante à era no arroz

The mutations are genetic changes in the genome sequences and have a significant role in biotechnology, genetics, and molecular biology even to find out the genome sequences of a cell DNA along with the viral RNA sequencing. The mutations are the alterations in DNA that may be natural or spontaneous and induced due to biochemical reactions or radiations which damage cell DNA. There is another cause of mutations which is known as transposons or jumping genes which can change their position in the genome during meiosis or DNA replication. The transposable elements can induce by self in the genome due to cellular and molecular mechanisms including hypermutation which caused the localization of transposable elements to move within the genome. The use of induced mutations for studying the mutagenesis in crop plants is very common as well as a promising method for screening crop plants with new and enhanced traits for the improvement of yield and production. The utilization of insertional mutations through transposons or jumping genes usually generates stable mutant alleles which are mostly tagged for the presence or absence of jumping genes or transposable elements. The transposable elements may be used for the identification of mutated genes in crop plants and even for the stable insertion of transposable elements in mutated crop plants. The guanine nucleotide-binding (GTP) proteins have an important role in inducing tolerance in rice plants to combat abiotic stress conditions.

Mutações são alterações genéticas nas sequências do genoma e têm papel significativo na biotecnologia, genética e biologia molecular, até mesmo para descobrir as sequências do genoma de um DNA celular junto com o sequenciamento do RNA viral. As mutações são alterações no DNA que podem ser naturais ou espontâneas e induzidas devido a reações bioquímicas ou radiações que danificam o DNA celular. Há outra causa de mutações, conhecida como transposons ou genes saltadores, que podem mudar sua posição no genoma durante a meiose ou a replicação do DNA. Os elementos transponíveis podem induzir por si próprios no genoma devido a mecanismos celulares e moleculares, incluindo hipermutação que causou a localização dos elementos transponíveis para se moverem dentro do genoma. O uso de mutações induzidas para estudar a mutagênese em plantas cultivadas é muito comum, bem como um método promissor para a triagem de plantas cultivadas com características novas e aprimoradas para a melhoria da produtividade e da produção. A utilização de mutações de inserção por meio de transposons ou genes saltadores geralmente gera alelos mutantes estáveis que são marcados quanto à presença ou ausência de genes saltadores ou elementos transponíveis. Os elementos transponíveis podem ser usados para a identificação de genes mutados em plantas de cultivo e até mesmo para a inserção estável de elementos transponíveis em plantas de cultivo mutadas. As proteínas de ligação ao nucleotídeo guanina (GTP) têm papel importante na indução de tolerância em plantas de arroz para combater as condições de estresse abiótico.

Era-like GTP protein gene expression in rice

Abstract The mutations are genetic changes in the genome sequences and have a significant role in biotechnology, genetics, and molecular biology even to find out the genome sequences of a cell DNA along with the viral RNA sequencing. The mutations are the alterations in DNA that may be natural or spontaneous and induced due to biochemical reactions or radiations which damage cell DNA. There is another cause of mutations which is known as transposons or jumping genes which can change their position in the genome during meiosis or DNA replication. The transposable elements can induce by self in the genome due to cellular and molecular mechanisms including hypermutation which caused the localization of transposable elements to move within the genome. The use of induced mutations for studying the mutagenesis in crop plants is very common as well as a promising method for screening crop plants with new and enhanced traits for the improvement of yield and production. The utilization of insertional mutations through transposons or jumping genes usually generates stable mutant alleles which are mostly tagged for the presence or absence of jumping genes or transposable elements. The transposable elements may be used for the identification of mutated genes in crop plants and even for the stable insertion of transposable elements in mutated crop plants. The guanine nucleotide-binding (GTP) proteins have an important role in inducing tolerance in rice plants to combat abiotic stress conditions.

Resumo Mutações são alterações genéticas nas sequências do genoma e têm papel significativo na biotecnologia, genética e biologia molecular, até mesmo para descobrir as sequências do genoma de um DNA celular junto com o sequenciamento do RNA viral. As mutações são alterações no DNA que podem ser naturais ou espontâneas e induzidas devido a reações bioquímicas ou radiações que danificam o DNA celular. Há outra causa de mutações, conhecida como transposons ou genes saltadores, que podem mudar sua posição no genoma durante a meiose ou a replicação do DNA. Os elementos transponíveis podem induzir por si próprios no genoma devido a mecanismos celulares e moleculares, incluindo hipermutação que causou a localização dos elementos transponíveis para se moverem dentro do genoma. O uso de mutações induzidas para estudar a mutagênese em plantas cultivadas é muito comum, bem como um método promissor para a triagem de plantas cultivadas com características novas e aprimoradas para a melhoria da produtividade e da produção. A utilização de mutações de inserção por meio de transposons ou genes saltadores geralmente gera alelos mutantes estáveis que são marcados quanto à presença ou ausência de genes saltadores ou elementos transponíveis. Os elementos transponíveis podem ser usados para a identificação de genes mutados em plantas de cultivo e até mesmo para a inserção estável de elementos transponíveis em plantas de cultivo mutadas. As proteínas de ligação ao nucleotídeo guanina (GTP) têm papel importante na indução de tolerância em plantas de arroz para combater as condições de estresse abiótico.

Era-like GTP protein gene expression in rice / Expressão do gene da proteína GTP semelhante à era no arroz

The mutations are genetic changes in the genome sequences and have a significant role in biotechnology, genetics, and molecular biology even to find out the genome sequences of a cell DNA along with the viral RNA sequencing. The mutations are the alterations in DNA that may be natural or spontaneous and induced due to biochemical reactions or radiations which damage cell DNA. There is another cause of mutations which is known as transposons or jumping genes which can change their position in the genome during meiosis or DNA replication. The transposable elements can induce by self in the genome due to cellular and molecular mechanisms including hypermutation which caused the localization of transposable elements to move within the genome. The use of induced mutations for studying the mutagenesis in crop plants is very common as well as a promising method for screening crop plants with new and enhanced traits for the improvement of yield and production. The utilization of insertional mutations through transposons or jumping genes usually generates stable mutant alleles which are mostly tagged for the presence or absence of jumping genes or transposable elements. The transposable elements may be used for the identification of mutated genes in crop plants and even for the stable insertion of transposable elements in mutated crop plants. The guanine nucleotide-binding (GTP) proteins have an important role in inducing tolerance in rice plants to combat abiotic stress conditions.

Mutações são alterações genéticas nas sequências do genoma e têm papel significativo na biotecnologia, genética e biologia molecular, até mesmo para descobrir as sequências do genoma de um DNA celular junto com o sequenciamento do RNA viral. As mutações são alterações no DNA que podem ser naturais ou espontâneas e induzidas devido a reações bioquímicas ou radiações que danificam o DNA celular. Há outra causa de mutações, conhecida como transposons ou genes saltadores, que podem mudar sua posição no genoma durante a meiose ou a replicação do DNA. Os elementos transponíveis podem induzir por si próprios no genoma devido a mecanismos celulares e moleculares, incluindo hipermutação que causou a localização dos elementos transponíveis para se moverem dentro do genoma. O uso de mutações induzidas para estudar a mutagênese em plantas cultivadas é muito comum, bem como um método promissor para a triagem de plantas cultivadas com características novas e aprimoradas para a melhoria da produtividade e da produção. A utilização de mutações de inserção por meio de transposons ou genes saltadores geralmente gera alelos mutantes estáveis que são marcados quanto à presença ou ausência de genes saltadores ou elementos transponíveis. Os elementos transponíveis podem ser usados para a identificação de genes mutados em plantas de cultivo e até mesmo para a inserção estável de elementos transponíveis em plantas de cultivo mutadas. As proteínas de ligação ao nucleotídeo guanina (GTP) têm papel importante na indução de tolerância em plantas de arroz para combater as condições de estresse abiótico.

The value of cell division cycle 42 in the efficacy and prognosis evaluation of arterial infusion chemotherapy for pancreatic cancer / 肿瘤研究与临床

Objective:To explore the value of cell division cycle 42 (CDC42) in the efficacy and prognosis evaluation of arterial infusion chemotherapy for pancreatic cancer.Methods:The clinical data of 100 patients with pancreatic cancer who underwent arterial infusion chemotherapy from January 2018 to January 2020 at Second People's Hospital of Wuhu were retrospectively analyzed, and all patients were divided into effective group (the complete remission and partial remission) and ineffective group (the stable disease and the progressive disease) according to the chemotherapy efficacy determined by CT. The clinicopathological characteristics of both groups were compared. The influencing factors of chemotherapy efficacy were determined by using multivariate logistic regression model analysis. The efficacy evaluated by CT examination was treated as the gold standard. The receiver operating characteristic (ROC) curve was used to analyze the value of CDC42 level predicting the efficacy of arterial infusion chemotherapy for pancreatic cancer patients before infusion chemotherapy. Survival analysis was performed by using Kaplan-Meier and log-rank test was also performed.Results:Among 100 patients with pancreatic cancer, there were 13 cases of complete remission, 30 cases of partial remission, 20 cases of stable disease, 37 cases of progressive disease; 43 cases in effective group and 57 cases in ineffective group. The proportions of tumor long diameter > 4 cm, TNM staging Ⅲ-Ⅳ, carcinoembryonic antigen (CA)199 > 37 U/ml, carcino-embryonic antigen (CEA) > 5 ng/ml, neutrophil-to-lymphocyte (NLR) > 2.8, serum total bilirubin > 34.2 μmol/L before infusion, CDC42 ≤ 1.11 μg/L, low differentiation degree and vascular invasion in ineffective group were higher than those in effective group (all P < 0.05). Tumor long diameter > 4 cm, TNM staging Ⅲ-Ⅳ, CA199 > 37 U/ml, CEA > 5 ng/ml before infusion, low differentiation degree, vascular invasion, and CDC42 ≤ 1.11 μg/L were independent risk factors for effectiveness of arterial infusion chemotherapy (all P < 0.05). The area under the ROC curve of CDC42 predicting the ineffectiveness of arterial infusion chemotherapy was 0.810 (95% CI 0.781-0.839, P <0.01), and the optimal cut-off value was 1.11 μg/L, the sensitivity was 96.25%, and the specificity was 63.13%. Survival curve analysis showed that the 2-year overall survival rate of patients with CDC42 > 1.11 μg/L was 58.93% which was greater than that of patients with CDC42 ≤ 1.11 μg/L (22.73%), and the difference was statistically significant ( χ2 = 14.99, P<0.001). Conclusion:CDC42 level is an independent influencing factor for the efficacy of arterial infusion chemotherapy in patients with pancreatic cancer, and it can effectively predict the prognosis of patients.

Relationship between long-term learning and memory impairment induced by sevoflurane anesthesia and PSD-95/Kalirin-7/Rac1 signaling pathway in neonatal rats / 中华麻醉学杂志

Objective:To evaluate the relationship between long-term learning and memory impairment induced by sevoflurane anesthesia and postsynaptic density protein-95 (PSD-95)/Kalirin-7/Ras-related C3 botulinum toxin substrate 1 (Rac1) signaling pathway in neonatal rats.Methods:Sixty SPF male Wistar rats, aged 7 days, weighing 12-18 g, were divided into 5 groups ( n=12 each) using a random number table method: control group (group C), 1% sevoflurane anesthesia for 2 h group (group S 1), 1% sevoflurane anesthesia for 4 h group (group S 2), 2% sevoflurane anesthesia for 2 h group (group S 3) and 2% sevoflurane anesthesia for 4 h group (group S 4). Morris water maze test was performed at 4, 8 and 12 weeks after anesthesia.The rats were sacrificed after the last Morris water maze test, and the hippocampal tissues were obtained for microscopic examination of the pathological changes (using HE staining), neuron apoptosis (by TUNEL staining), and expression of PSD-95, Kalirin-7 and Rac1 protein and mRNA (by Western blot and quantitative real-time polymerase chain reaction). The apoptosis rate was calculated. Results:Compared with group C, the escape latency was significantly prolonged, the number of crossing the original platform was reduced, the time of stay in the target quadrant was shortened, and the apoptosis rate of hippocampal neurons was increased at 4th, 8th and 12th weeks after anesthesia, phosphorylated Rac1/Rac1 ratio was decreased, and the expression of PSD-95 and Kalirin-7 protein and mRNA was down-regulated in S 1, S 2, S 3 and S 4 groups ( P<0.05). Compared with group S 4, the escape latency was significantly shortened, the number of crossing the original platform was increased, the time of stay in the target quadrant was prolonged, and the apoptosis rate of hippocampal neurons was decreased, phosphorylated Rac1/Rac1 ratio was increased, the expression of PSD-95 and Kalirin-7 protein and mRNA was up-regulated, and the histopathological changes of hippocampal tissues were attenuated in S 1, S 2 and S 3 groups ( P<0.05). Conclusions:The mechanism by which sevoflurane anesthesia induces long-term learning and memory impairment may be related to inhibition of activity of PSD-95/Kalirin-7/Rac1 signaling pathway in hippocampi of neonatal rats.

Effects of GNG4 on DNA damage repair and chemosensitivity of ovarian cancer cisplatin-resistant A2780/DDP cells / 肿瘤研究与临床

Objective:To investigate the correlation of GNG4 with DNA damage repair and chemosensitivity of ovarian cancer cisplatin-resistant A2780/DDP cells.Methods:A2780/DDP cells were divided into 500 ng/ml cisplatin group (cDDP group), short hairpin RNA (shRNA)-GNG4 silencing GNG4 expression group (shRNA group), 500 ng/ml cisplatin and shRNA-GNG4 intervention group (shRNA+cDDP group), and non cisplatin and shRNA-GNG4 intervention group (blank control group). Western blot was used to detect the expressions of GNG4 and γH2AX proteins in each group; DNA damage in each group was detected by single cell gel electrophoresis. The focus formation of γH2AX gene at the injury site was detected by immunofluorescence. The ability of cell clone formation was detected by plate clone formation experiment.Results:Compared with the other three groups, the expression level of GNG4 protein in shRNA+cDDP group was the lowest, the expression level of γH2AX protein was the highest, and the differences were statistically significant (all P < 0.01). Single cell gel electrophoresis assay showed that the comet tail DNA% in blank control group, cDDP group, shRNA group and shRNA+cDDP group were (7.7±2.5)%, (12.3±3.6)%, (20.1±2.1)%, (38.6±2.8)%, respectively, and Olive trailing distance were 5.12±1.89, 8.23±2.97, 14.99±3.65, 22.43±3.17, respectively, the comet tail DNA% and Olive tail distance in shRNA+cDDP group were higher than those in the other three groups, and the differences were statistically significant (all P < 0.05). Immunofluorescence assay showed that the focus numbers of γH2AX in each cell of blank control group, cDDP group, shRNA group and shRNA+cDDP group were 4.2±0.7, 5.1±0.5, 26.8±3.3, 71.3±6.2, respectively, the shRNA+cDDP group was higher than the other three groups, and the differences were statistically significant (all P < 0.05). The clone formation rates of blank control group, cDDP group, shRNA group and shRNA+cDDP group were (78.27±5.01)%, (45.67±3.29)%, (26.20±5.76)%, (1.56±0.21)%, respectively, the shRNA+cDDP group was lower than the other three groups, and the differences were statistically significant (all P < 0.001). Conclusions:Down-regulation of GNG4 expression can increase the cisplatin sensitivity of ovarian cancer A2780/DDP cells, which may be achieved by inhibiting the DNA damage repair function induced by cisplatin.

Non-invasive prenatal test for autosomal dominant Charcot-Marie-Tooth disease in a fetus using droplet digital polymerase chain reaction / 中华围产医学杂志

We report the use of droplet digital polymerase chain reaction (ddPCR) in non-invasive prenatal test fetal Charcot-Marie-Tooth disease (CMT) caused by MFN2 gene mutation. The proband, namely the husband, was found with heterozygous mutation of c.919A>G(p.K307E) in the MFN2 gene, which was diagnosed as CMT type 2A2A at a local hospital. The proband's wife underwent genetic counseling after conception at the First Affiliated Hospital of Zhengzhou University. Peripheral blood obtained from the pregnant woman was analyzed by ddPCR at eight gestational weeks, which found the fetus to carry a paternal pathogenic gene mutation. Sanger sequencing for the chorionic sample at 11 gestational weeks further verified that the fetus was a c.919G>A(p.K307E) heterozygous mutation carrier, the same as the proband. ddPCR could be applied to cell-free fetal DNA to detect the paternal pathogenic gene mutation in the non-invasive prenatal test.

GTP Cyclohydrolase Functions in Fish Color Formation / 中国生物化学与分子生物学报

Guanosine triphosphate cyclohydrolase (GTP cyclohydrolase,Gch) is a protease with a GTP- cyclohydro domain, which is widely found in vertebrates and invertebrates.Mammals and birds only have Gch 1.In teleost and amphibian, other two paralogs (Gch2 and Gch3) also exists besides Gchl, which also displayed functional differences.Gch is a rate-limiting enzyme that ultimately synthesized the tetrahydrobiopterin (BH4) using guanosine triphosphate as a substrate.BH4 is an essential coenzyme of aromatic amino acid hydroxylase and contributes to the synthesis of various hormones and neurotransmitters.The Gch is an initial step in the catalysis of various pterin biosynthesis and plays important roles in a series of physiological and pathological processes, such as skin pigmentation, ocular pigmentation, methotrexate, folic acid, and tetrahydrobiopterin.The physiological function of Gch is inextricably linked to the biosynthesis of BH4.As the only rate-limiting enzyme in BH4 biosynthesis, the activity of Gch is a useful indicator for the development of neurons and pigment cells.Besides, it is also an important marker of pigment synthesis and neurotransmitter biosynthesis.Nowadays, the functions of Geh in pathogenesis of tumor and cardiovascular diseases have been widely concerned, while the researches on the pigmentation and color formation are mainly concentrated in insects, and rarely in teleost.Therefore, this article summarized the characteristics of Gch genes, protein and the functions of Gch in fish coloration, which has important guiding significance for further illustration the mechanism of Gch in teleost pigmentation and fish color genetic improvements.

Identification of phospholipase C β downstream effect on transient receptor potential canonical 1/4, transient receptor potential canonical 1/5 channels

Gα(q)-coupled receptor stimulation was implied in the activation process of transient receptor potential canonical (TRPC)1/4 and TRPC1/5 heterotetrameric channels. The inactivation occurs due to phosphatidylinositol 4,5-biphosphate (PI(4,5)P₂) depletion. When PI(4,5)P₂ depletion was induced by muscarinic stimulation or inositol polyphosphate 5-phosphatase (Inp54p), however, the inactivation by muscarinic stimulation was greater compared to that by Inp54p. The aim of this study was to investigate the complete inactivation mechanism of the heteromeric channels upon Gα(q)-phospholipase C β (Gα(q)-PLCβ) activation. We evaluated the activity of heteromeric channels with electrophysiological recording in HEK293 cells expressing TRPC channels. TRPC1/4 and TRPC1/5 heteromers undergo further inhibition in PLCβ activation and calcium/protein kinase C (PKC) signaling. Nevertheless, the key factors differ. For TRPC1/4, the inactivation process was facilitated by Ca²⁺ release from the endoplasmic reticulum, and for TRPC1/5, activation of PKC was concerned mostly. We conclude that the subsequent increase in cytoplasmic Ca²⁺ due to Ca²⁺ release from the endoplasmic reticulum and activation of PKC resulted in a second phase of channel inhibition following PI(4,5)P₂ depletion.

Relationship between Rho GTPase family regulation and vascular endothelial barrier function / 药学学报

Injury of vascular endothelial barrier function is implicated in several pathophysiological processes. The integrity of vascular endothelium is regulated by cytoskeleton and cell-cell junctions. Small guanosine triphosphatases of the Rho family (Rho GTPases) are known to play a central role in vascular endothelial barrier function. It has been reported that RhoA, Rac1, Cdc42 and RhoB are involved and they exert both positive and negative effect on endothelial barrier integrity, depending on their subcellular location. When inflammatory factors such as thrombin attack the vascular endothelial cells, GEF of RhoA will be widely distributed throughout the cells. Thus, activated RhoA causes aggregation of F-actin fibers in a short time and disrupts the vascular endothelial barrier, a process named acute cell contraction. However, RhoA may also induce the production and maturation of intercellular junctions in new cells. Rac1 and Cdc42 help to maintain the integrity of vascular endothelial barrier at the resting state. They cause the phosphorylation of LIM kinase and inhabitation of cofilin, resulting in less remodeling of cytoskeletal in the vascular endothelial cells. On the other hand, Cdc42 can translocate to the cortex rapidly after a stimulation, where Cdc42 will activate the myosin Ⅱ and promote the reorganization of adjective junction to facilitate the recovery of vascular endothelial barrier. In this review, we overviewed how Rho GTPases regulate the vascular endothelial barrier integrity.

Gamma-glutamyl transpeptidase (γ-GTP) has an ambivalent association with hypertension and atherosclerosis among elderly Japanese men: a cross-sectional study

BACKGROUND@#Even though there is bidirectional association between hypertension and atherosclerosis, atherosclerosis itself is involved in the process of endothelial repair. To clarify the association of endothelial repair with hypertension, a cross-sectional study was conducted.@*METHODS@#We conducted a cross-sectional study of 562 elderly Japanese men aged 60-69. As gamma-glutamyl transpeptidase (γ-GTP) could act as a marker of oxidative stress that injures endothelial cell and higher levels of CD34-positive cell indicate a higher activity of endothelial repair, we therefore performed a CD34-positive level specific analysis of γ-GTP on atherosclerosis and hypertension.@*RESULTS@#In the present study population, hypertension was independently and positively associated with atherosclerosis (multivariable odds ratio (OR) = 2.09 (1.30, 3.35)). Among participants with high CD34-positive cells, γ-GTP showed significant and positive association with atherosclerosis (OR of the log-transformed value of γ-GTP (OR) = 2.26 (1.32, 3.86)) but not with hypertension (OR = 0.77 (0.51, 1.17)). Among participants with low CD34-positive cells, even γ-GTP showed no significant association with atherosclerosis (OR = 0.92 (0.51, 1.68)), but was significantly and positively associated with hypertension (OR = 1.99 (1.27, 3.12)).@*CONCLUSIONS@#γ-GTP revealed to have ambivalent association with hypertension and atherosclerosis. Active endothelial repair that is associated with atherosclerosis might have beneficial association with hypertension.

Progress on study of guanosine triphosphate cyclohydrolase 1 / 中国医师杂志

Guanosine triphosphate cyclohydrolase 1 (GTPCH1) is a protein encoded by the GCH1 gene,which catalyze GTP to tetrahydrofolinine (BH4) under physiological condition.BH4 is a coenzyme of aromatic amino acid hydroxylase and a cofactor of nitric oxide synthases.BH4 involves in the synthesis of various hormones and neurotransmitters and plays an important role in a series of pathophysiological processes in vivo.Recent studies showed that GTPCH1 is involved in the pathogenesis of neuropathic pain,doparesponsive dystonia,cancer and cardiovascular diseases.In this review,we will discuss the role of GTPCH1 in those diseases mentioned above.

Research advances of the relationship between the GTP cyclohydrolase 1, tetrahydrobiopterin and neuropathic pain / 中国医师进修杂志

Neuropathic pain is one of the most common chronic pain in clinic, and its treatment has always been a global difficulty, mainly because its pathogenesis is unknown. GTP cyclohydrolase 1 (GCH1) mediated neuropathic pain by regulating inflammatory cytokines. Besides, GCH1 is also the main speed limit in the process of tetrahydrobiopterin (BH4) enzyme synthesis. BH4 is a necessary cofactor for the synthesis of various inflammatory factors and neurotransmitters. In the study of its relationship with pain, it was found that when the GCH1 gene was silent, the pain was relieved and the expression of BH4 decreased. The results suggest that GCH1 and BH4 have a certain relationship with neuropathic pain.

Expression of Rap1, guanosine triphosphate Rap1, vascular endothelial growth factor and β-catenin in experimental choroidal neovascularization / 中华眼底病杂志

Objective To observe the expression ofRapl,guanosine triphosphate-Rapl (GTP-Rapl),vascular endothelial growth factor (VEGF) and β-catenin in experimental choroidal neovascularization (CNV).Methods Forty-two brown Norwegian rats were randomly divided into a blank control group (7 rats) and a model group (35 rats).Both eyes were enrolled.The CNV model was established by holmium ion laser photocoagulation in the model group.At 3,7,14,21,and 28 days after photocoagulation,fluorescein fundus angiography (FFA) and choroidal vascular smear were performed to observe the degree of fluorescein leakage and CNV area in rats;Western blot and real-time quantitative polymerase chain reaction (RT-PCR) were used to detect the expression ofRap1,GTP-Rap1,VEGF,β-catenin and mRNA in CNV.Results The results of FFA examination showed that a large disc-shaped fluorescein leaked in the photo-condensation spot 14 days after photocoagulation.Laser confocal microscopy showed that compared with 7 days after photocoagulation,CNV area increased at 14,21,28 days after photocoagulation,and the difference were statistically significant (t=3.725,5.532,3.605;P＜0.05).Western blot showed that there was no significant difference in the relative expression of Rap1 protein in CNV at different time points after photocoagulation between the two groups (P=0.156).Compared with the blank control group,the relative expression of GTP-Rap1 protein was significantlydecreased,the relative expression of VEGF and β-catenin protein were significantly increased in the model group (P=0.000).The results of RT-PCR showed that there was no significant difference in the relative expression of Rap 1 mRNA at different time points after photocoagulation between the two groups (P=0.645),but there were significant difference in the relative expression of β-catenin mRNA (P=0.000).At 7,14,21 and 28 days after photocoagulation,there were significant difference in the relative expression of GTP-Rap 1 and VEGF mRNA between the two groups (P=0.000).Conclusions The expression of GTP-Rap1 in experimental CNV is significantly lower than that in normal rats.

Investigation of protective mechanisms of Atorvastatin against high glucose environment-induced injuries of myocardial microvascular endothelial cells / 中华老年医学杂志

Objective To investigate the protective mechanisms of Atorvastatin against high glucose environment-induced injuries of myocardial microvascular endothelial cells. Methods Myocardial microvascular endothelial cells(MMECs)in SD rat were cultured and divided into groups of control group ,hyperglycemia group ,atorvastatin group ,and atorvastatin + high glucose group. The level of reactive oxygen species (ROS)was assayed using Superoxide Assay Kit. Apoptosis of cells was detected by terminal deoxynucleotidyl transferase(TdT)-mediated dUTP nick end labeling(TUNEL) . The expression levels of Akt1 and β1-Integrin were assayed by short-interfering RNA (siRNA ) technique ,and the levels of small GTP-binding protein dissociation stimulator (SmgGDS) expression were measured using Western blot. Results (1)The level of ROS was higher in the high glucose group than in the control group(t=4.154 ,P <0.01) ,and lower in both Atorvastatin group and the Atorvastatin + high glucose group than in the high glucose group (t= 4.233 and 2.893 ,both P <0.05). (2)The proportion of apoptotic cells was higher in the high glucose group than in the control group(t= 4.058 ,P < 0.01) ,and lower in both Atorvastatin group and the Atorvastatin + high glucose group than in the high glucose group(t=4.157 and 2.601 ,both P<0.05).(3)The expression level of Akt1 was lower in the high glucose group and the high glucose + Atorvastatin group than in the mock control group after transfection of Akt1-siRNA(t=4.058 and 4.167 ,both P<0.01).The expression level of β1-integrin was lower in the high glucose group and the high glucose + atorvastatin group than in the mock control group after transfection of β1-integrin-siRNA (t=4.073 and 4.215 , both P<0.01). (4)Western blot analysis showed the following results. First ,the relative expression levels of SmgGDS in both the low dose(1 μmol/L)and high dose(10 μmol/L)of atorvastatin group were higher than in the control group (t= 2.671 and 2.832 ,both P < 0.05).Second ,the relative expression level of SmgGDS in the high dose group were higher than in the low dose group (t=2.612 , P< 0.05 ). Third ,after transfection of Akt1-siRNA ,the expression level of SmgGDS in the high glucose + Atorvastatin group and the high glucose group was decreased ;and the level was higher in the high glucose + atorvastatin + mock group than in the high glucose + mock group(t=4.051 ,P<0.01).Fourth ,after transfection of β1-integrin-siRNA ,the expression level of SmgGDS was lower in high glucose + Atorvastatin group and the high glucose group than in the high glucose +Atorvastatin + mock group ;the level was higher in the high glucose + Atorvastatin + mock group than in the high glucose + mock group(t= 4.068 ,P < 0.01).Fifth ,the expression level of Akt phosphorylation in the high glucose group and the high glucose + Atorvastatin group was higher at 10 minutes than at five minutes(t=2.608 ,P<0.05) ,and higher at 15 minutes than at 10 minutes(t=3.127 ,P <0.05). After transfection of β1-integrin-siRNA ,the expression level of p-Akt /t-Akt was lower in the high glucose group than in the high glucose + mock group(t= 3.371 ,P < 0.05). Conclusions Atorvastatin treatment protects myocardial microvascular endothelial cells possibly by up-regulating SmgGDS through β1-integrin/Akt1 against high glucose environment-induced oxidative stress and apoptosis injuries.

ω-3 polyunsaturated fatty acids inhibits RHO-ROCK1 signaling pathway in SGC7901 gastric cancer cells / 中国医师杂志

Objective This study was aimed to investigate the effects of ω-3 polyunstaurated fatty acids (ω-3 PUFAs) on the growth of gastric cancer cells in nude mice,and to find whether the Ros homolog gene Rho-associated coiled-coil containing protein kinase 1 (RHO-ROCK1) signaling pathway is involved.Methods 16 BALB/C nude mice were injected subcutaneously with SGC7901 gastric cancer cells to establish the tumor-bearing mouse model.The mice were randomized:control group (normal saline) and intervention group (ω-3 PUFAs).The mRNA expression of Ros homolog gene family,member A (RHOA),RHOC,and ROCK1 in tumor tissue were detected by quantitative polymerase chain reaction (qPCR).Immunofluorescence and Western blot were used to detect RHOA,RHOC,and ROCK1 protein expression.Results The volume and weight of the tumors in the ω-3 PUFAs group were slightly smaller than that in the control group (P ＞ 0.05).Compared to the control group,hematoxylin and eosin staining showed multifocal tumor necrosis in the ω-3 PUFAs group,while the tumors of the control group showed abundant blood supply.qPCR and Western blot showed that the mRNA and proteins expression of RHOA and ROCK1 in the ω-3 PUFAs group was significantly lower than those in the control group (P ＜ 0.05).The immunofluorescence redults also showed that the expression of these proteins in the ω-3 PUFAs group was slightly lower than that in the control group.Conclusions These results suggested that ω-3 PUFAs may affect the growth of gastric cancer in nude mice by affecting the expression of RHOA,RHOC and ROCK1,thus inhibiting the excessive proliferation of gastric cancer cells and leading to tumor necrosis.