S100 proteins and their implication in the thalamus: an integrative review / Las proteínas S100 y su implicación en el tálamo: una revisión integradora

SUMMARY: S100 proteins belong group of calcium-binding proteins and are present in physiological intracellular and extracellular regulatory activities, such as cell differentiation, and act in inflammatory and neoplastic pathological processes. Recently, its expressions in the nervous system have been extensively studied, seeking to elucidate its action at the level of the thalamus: A structure of the central nervous system that is part of important circuits, such as somatosensory, behavioral, memory and cognitive, as well as being responsible for the transmission and regulation of information to the cerebral cortex. This article is an integrative review of scientific literature, which analyzed 12 studies present in Pubmed. The analysis showed that the relationship of S100 proteins and the thalamus has been described in neoplastic processes, mental disorders, hypoxia, trauma, stress, infection, Parkinson's disease and epilepsy. In summary, it is possible to conclude that this protein family is relevant as a marker in processes of thalamic injury, requiring further studies to better understand its clinical, preclinical meanings and its prognostic value.

Las proteínas S100 pertenecen al grupo de proteínas fijadoras de calcio y están presentes en actividades reguladoras fisiológicas intracelulares y extracelulares, como la diferenciación celular, y actúan en procesos patológicos inflamatorios y neoplásicos. Recientemente, sus expresiones en el sistema nervioso han sido ampliamente estudiadas, buscando dilucidar su acción a nivel del tálamo: una estructura del sistema nervioso central que forma parte de importantes circuitos, como el somatosensorial, conductual, de memoria y cognitivo, así como además de ser responsable de la transmisión y regulación de la información a la corteza cerebral. Este artículo es una revisión integradora de la literatura científica, que analizó 12 estudios presentes en Pubmed. El análisis mostró que la relación de las proteínas S100 y el tálamo ha sido descrita en procesos neoplásicos, trastornos mentales, hipoxia, trauma, estrés, infección, enfermedad de Parkinson y epilepsia. En resumen, es posible concluir que esta familia de proteínas es relevante como marcador en procesos de lesión talámica, requiriendo más estudios para comprender mejor su significado clínico, preclínico y su valor pronóstico.

NKD1 promotes glucose uptake in colon cancer cells by activating YWHAE transcription / 南方医科大学学报

OBJECTIVE@#Bo investigate the regulatory relationship between NKD1 and YWHAE and the mechanism of NKD1 for promoting tumor cell proliferation.@*METHODS@#HCT116 cells transfected with pcDNA3.0-NKD1 plasmid, SW620 cells transfected with NKD1 siRNA, HCT116 cells with stable NKD1 overexpression (HCT116-NKD1 cells), SW620 cells with nkd1knockout (SW620-nkd1-/- cells), and SW620-nkd1-/- cells transfected with pcDNA3.0-YWHAE plasmid were examined for changes in mRNA and protein expression levels of YWHAE using qRT-PCR and Western blotting. Chromatin immunoprecipitation (ChIP) assay was used to detect the binding of NKD1 to the promoter region of YWHAE gene. The regulatory effect of NKD1 on YWHAE gene promoter activity was analyzed by dual-luciferase reporter gene assay, and the interaction between NKD1 and YWHAE was analyzed with immunofluorescence assay. The regulatory effect of NKD1 on glucose uptake was examined in the tumor cells.@*RESULTS@#In HCT116 cells, overexpression of NKD1 significantly enhanced the expression of YWHAE at both the mRNA and protein levels, while NKD1 knockout decreased its expression in SW620 cells (P < 0.001). ChIP assay showed that NKD1 protein was capable of binding to the YWHAE promoter sequence; dual luciferase reporter gene assay showed that NKD1 overexpression (or knockdown) in the colon cancer cells significantly enhanced (or reduced) the transcriptional activity of YWHAE promoter (P < 0.05). Immunofluorescence assay demonstrated the binding of NKD1 and YWHAE proteins in colon cancer cells. NKD1 knockout significantly reduced glucose uptake in colon cancer cells (P < 0.01), while YWHAE overexpression restored the glucose uptake in NKD1-knockout cells (P < 0.05).@*CONCLUSION@#NKD1 protein activates the transcriptional activity of YWHAE gene to promote glucose uptake in colon cancer cells.

Mechanism of n-butanol alcohol extract of Baitouweng Decoction in treatment of vulvovaginal candidiasis based on negative regulation of NLRP3 inflammasome via PKCδ/NLRC4/IL-1Ra axis / 中国中药杂志

This study aimed to explore the mechanism of n-butanol alcohol extract of Baitouweng Decoction(BAEB) in the treatment of vulvovaginal candidiasis(VVC) in mice based on the negative regulation of NLRP3 inflammasome via PKCδ/NLRC4/IL-1Ra axis. In the experiment, female C57BL/6 mice were divided randomly into the following six groups: a blank control group, a VVC model group, high-, medium-, and low-dose BAEB groups(80, 40, and 20 mg·kg~(-1)), and a fluconazole group(20 mg·kg~(-1)). The VVC model was induced in mice except for those in the blank control group by the estrogen dependence method. After modeling, no treatment was carried out in the blank control group. The mice in the high-, medium-, and low-dose BAEB groups were treated with BAEB at 80, 40, and 20 mg·kg~(-1), respectively, and those in the fluconazole group were treated with fluconazole at 20 mg·kg~(-1). The mice in the VVC model group received the same volume of normal saline. The general state and body weight of mice in each group were observed every day, and the morphological changes of Candida albicans in the vaginal lavage of mice were examined by Gram staining. The fungal load in the vaginal lavage of mice was detected by microdilution assay. After the mice were killed, the degree of neutrophil infiltration in the vaginal lavage was detected by Papanicolaou staining. The content of inflammatory cytokines interleukin(IL)-1β, IL-18, and lactate dehydrogenase(LDH) in the vaginal lavage was tested by enzyme-linked immunosorbent assay(ELISA), and vaginal histopathology was analyzed by hematoxylin-eosin(HE) staining. The expression and distribution of NLRP3, PKCδ, pNLRC4, and IL-1Ra in vaginal tissues were measured by immunohistochemistry(IHC), and the expression and distribution of pNLRC4 and IL-1Ra in vaginal tissues were detected by immunofluorescence(IF). The protein expression of NLRP3, PKCδ, pNLRC4, and IL-1Ra was detected by Western blot(WB), and the mRNA expression of NLRP3, PKCδ, pNLRC4, and IL-1Ra was detected by qRT-PCR. The results showed that compared with the blank control group, the VVC model group showed redness, edema, and white secretions in the vagina. Compared with the VVC model group, the BAEB groups showed improved general state of VVC mice. As revealed by Gram staining, Papanicolaou staining, microdilution assay, and HE staining, compared with the blank control group, the VVC model group showed a large number of hyphae, neutrophils infiltration, and increased fungal load in the vaginal lavage, destroyed vaginal mucosa, and infiltration of a large number of inflammatory cells. BAEB could reduce the transformation of C. albicans from yeast to hyphae. High-dose BAEB could significantly reduce neutrophil infiltration and fungal load. Low-and medium-dose BAEB could reduce the da-mage to the vaginal tissue, while high-dose BAEB could restore the damaged vaginal tissues to normal levels. ELISA results showed that the content of inflammatory cytokines IL-1β, IL-18, and LDH in the VVC model group significantly increased compared with that in the blank control group, and the content of IL-1β, IL-18 and LDH in the medium-and high-dose BAEB groups was significantly reduced compared with that in the VVC model group. WB and qRT-PCR results showed that compared with the blank control group, the VVC model group showed reduced protein and mRNA expression of PKCδ, pNLRC4, and IL-1Ra in vaginal tissues of mice and increased protein and mRNA expression of NLRP3. Compared with the VVC model group, the medium-and high-dose BAEB groups showed up-regulated protein and mRNA expression of PKCδ, pNLRC4, and IL-1Ra in vaginal tissues and inhibited protein and mRNA expression of NLRP3 in vaginal tissues. This study indicated that the therapeutic effect of BAEB on VVC mice was presumably related to the negative regulation of NLRP3 inflammasome by promoting PKCδ/NLRC4/IL-1Ra axis.

PDCD6 Promotes Hepatocellular Carcinoma Cell Proliferation and Metastasis through the AKT/GSK3β/β-catenin Pathway / 生物医学与环境科学（英文）

OBJECTIVE@#Programmed cell death 6 (PDCD6), a Ca 2+-binding protein, has been reported to be aberrantly expressed in all kinds of tumors. The aim of this study was to explore the role and mechanism of PDCD6 in hepatocellular carcinomas (HCCs).@*METHODS@#The expression levels of PDCD6 in liver cancer patients and HCC cell lines were analyzed using bioinformatics and Western blotting. Cell viability and metastasis were determined by methylthiazol tetrazolium (MTT) and transwell assays, respectively. And Western blotting was used to test related biomarkers and molecular pathway factors in HCC cell lines. LY294002, a PI3K inhibitor inhibiting AKT, was used to suppress the AKT/GSK3β/β-catenin pathway to help evaluate the role of this pathway in the HCC carcinogenesis associated with PDCD6.@*RESULTS@#The analysis of The Cancer Genome Atlas Database suggested that high PDCD6 expression levels were relevant to liver cancer progression. This was consistent with our finding of higher levels of PDCD6 expression in HCC cell lines than in normal hepatocyte cell lines. The results of MTT, transwell migration, and Western blotting assays revealed that overexpression of PDCD6 positively regulated HCC cell proliferation, migration, and invasion. Conversely, the upregulation of PDCD6 expression in the presence of an AKT inhibitor inhibited HCC cell proliferation, migration, and invasion. In addition, PDCD6 promoted HCC cell migration and invasion by epithelial-mesenchymal transition. The mechanistic investigation proved that PDCD6 acted as a tumor promoter in HCC through the AKT/GSK3β/β-catenin pathway, increasing the expression of transcription factors and cellular proliferation and metastasis.@*CONCLUSION@#PDCD6 has a tumor stimulative role in HCC mediated by AKT/GSK3β/β-catenin signaling and might be a potential target for HCC progression.

Analysis of SLC25A13 gene variants in 16 infants with intrahepatic cholestasis caused by citrin protein deficiency / 中华医学遗传学杂志

OBJECTIVE@#To explore the characteristics of SLC25A13 gene variants in 16 infants with neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD).@*METHODS@#The infants were subjected to high-throughput DNA sequencing for coding exons and flanking regions of the target genes. Suspected variants were verified by Sanger sequencing and bioinformatic analysis.@*RESULTS@#Among the 16 NICCD cases, 15 were found to harbor pathogenic variants. Among these, IVS14-9A>G, c.1640G>A, c.762T>A, c.736delG, c.1098Tdel and c.851G>A were previously unreported.@*CONCLUSION@#Six novel SLC25A13 variants were found by high-throughput sequencing, which has enriched the spectrum of SLC25A13 gene variants and provided a basis for genetic counseling and prenatal diagnosis.

Immunohistochemical distribution of calcium binding proteins in the cochlear nuclear complex of circling mice / Distribución inmunohistoquímica de proteínas de unión a calcio en el complejo nuclear coclear de ratones circulantes

SUMMARY: The term "circling mouse" refers to an animal model of deafness, in which the mouse exhibits circling, head tossing, and hyperactivity, with pathological features including degenerated spiral ganglion cells in the cochlea, and the loss of the organ of Corti. The cochlear nuclear (CN) complex, a part of the auditory brain circuit, is essential to process both ascending and descending auditory information. Considering calcium's (Ca2+) importance in homeostasis of numerous biological processes, hearing loss by cochlear damage, either by ablation or genetic defect, could cause changes in the Ca2+ concentration that might trigger functional and structural alterations in the auditory circuit. However, little is known about the correlation of the central nervous system (CNS) pathology in circling mice, especially of the auditory pathway circuit and Ca2+ changes. This present study investigates the distribution of Ca2+- binding proteins (CaBPs), calbindin D-28k (CB), parvalbumin (PV), and calretinin (CR) by using a free floating immunohistochemical method inthe CN of the wild-type mouse (+/+), the heterozygous mouse (+/cir), and the homozygous (cir/cir) mouse. CaBPs are well known to be an important factor that regulates Ca2+ concentrations. Compared with the dorsal and ventral cochlear nuclei of +/+ and +/ cirmice, prominent decreases of CaBPs' immunoreactivity (IR) in cir/cirmice were observed in the somas, as well as in the neuropil. The present study reportson the overall distribution and changes in the immunoreactivity of CaBPs in the CN of cir/cirmice because ofa hearing defect. This data might be helpful to morphologically elucidate CNS disorders and their relation to CaBPs immunoreactivity related to hearing defects.

RESUMEN: El término "ratón circulante" se refiere a un modelo animal con sordera, en el que el ratón exhibe hiperactividad, movimientos circulares y movimientos de la cabeza, con características patológicas que incluyen células ganglionares espirales degeneradas en la cóclea, un canal de Rosenthal vacío y la pérdida del órgano de Corti. El complejo nuclear coclear (CN), una parte del circuito cerebral auditivo, es esencial para procesar la información auditiva tanto ascendente como descendente. Considerando la importancia del calcio (Ca2+) en la homeostasis de numerosos procesos biológicos, la hipoacusia por daño coclear, por ablación o por defecto genético, podría provocar cambios en la concentración de Ca2+que pueden desencadenar alteraciones funcionales y estructurales en el circuitoauditivo. Sin embargo, existe poca información de la correlación de la patología del sistema nervioso central (SNC) en ratones circulantes, especialmente del circuito de la víaauditiva y los cambios de Ca2+. Este estudio nvestiga la distribución de proteínas de unión a Ca2+ (CaBP), calbindina D-28k (CB), parvalbúmina (PV) y calretinina (CR) mediante el uso de un método inmunohistoquímico de flotaciónlibre en el CN del ratón de tiposalvaje (+/+), el ratón heterocigoto (+/cir) y el ratón homocigoto (cir/cir). Se sabe que los CaBP son un factor importante que regula las concentraciones de Ca2+. En comparación con los núcleos cocleares dorsal y ventral de los ratones +/+ y +/ cir, se observaron disminuciones prominentes de la inmunorreactividad (IR) de CaBPs en los ratonescir/cir en los somas, asícomo en el neuropilo. El presente estudio informa sobre la distribución general y los cambios en la inmunorreactividad de CaBP en el CN de ratones cir/cir debido a un defecto auditivo. Estos datos podrían ser útiles para dilucidar morfológicamente los trastornos del SNC y su relación con la inmunorreactividad de CaBP relacionada con los defectosauditivos.

An epidermal growth factor motif of developmental endothelial locus 1 protein inhibits efficient angiogenesis in explanted squamous cell carcinoma in vivo

ABSTRACT Background: Cancer gene therapy using a nonviral vector is expected to be repeatable, safe, and inexpensive, and to have long-term effectiveness. Gene therapy using the E3 and C1 (E3C1) domain of developmental endothelial locus-1 (Del1) has been shown to improve prognosis in a mouse transplanted tumor model. Objective: In this study, we examined how this treatment affects angiogenesis in mouse transplanted tumors. Materials and methods: Mouse transplanted tumors (SCCKN human squamous carcinoma cell line) were injected locally with a nonviral plasmid vector encoding E3C1 weekly. Histochemical analysis of the transplanted tumors was then performed to assess the effects of E3C1 on prognosis. Results: All mice in the control group had died or reached an endpoint within 39 days. In contrast, one of ten mice in the E3C1 group had died by day 39, and eight of ten had died or reached an endpoint by day 120 (p < 0.01). Enhanced apoptosis in tumor stroma was seen on histochemical analyses, as was inhibited tumor angiogenesis in E3C1-treated mice. In addition, western blot analysis showed decreases in active Notch and HEY1 proteins. Conclusion: These findings indicate that cancer gene therapy using a nonviral vector encoding E3C1 significantly improved life-span by inhibiting tumor angiogenesis. (REV INVEST CLIN. 2021;73(1):39-51)

Construction of a Stable Expression Cell Line of Human Phospholamban / 法医学杂志

OBJECTIVES@#To construct a cell line that can stably express human phospholamban(PLN) and initially explore its application in the study of myocardial toxicity mechanism.@*METHODS@#FastCloning method was used to insert the open reading frame sequence of target gene PLN into eukaryotic expression vector pcDNA5/FRT/TO(hereinafter referred to as pDFT) to construct the pDFT-PLN-Flag plasmid. The Flp-InTM T-RExTM 293 cells were generated by cotransfection of the constructed plasmid and pOG44 plasmid to express the target gene. Successfully recombined monoclonal cell lines were screened by hygromycin B resistance. Western blot and indirect immunofluorescence (IFA) were used to examine the expression of the target protein in recombinant cells. After the cell line was exposed to aconitine, it was verified by Western blot to detect changes in PLN protein phosphorylation.@*RESULTS@#After PCR amplification of the recombinant plasmid and DNA electrophoresis, the length of the amplified product is the same as the known PLN gene fragment, which is consistent with the open reading frame (ORF) sequence of the human PLN gene after sequencing. IFA and Western blot showed that the constructed proliferation cell line can stably express high levels of human PLN under induction and regulation. Preliminary results showed that the phosphorylation level of Thr17-PLN decreased after two hours of exposure to 1 μmol/L aconitine.@*CONCLUSIONS@#This human cell line can stably express PLN and can be used to study the mechanism of action of aconitine on the cell at molecular level.

Preliminary study on mechanism of electroacupuncture with the involvement of SERCA2a/PLB on the synergistic and attenuated effect of aconitine in treatment of heart failure / 中国针灸

OBJECTIVE@#To investigate the mechanism of electroacupuncture (EA) with the involvement of sarcoplasmic reticulum Ca@*METHODS@#Thirty SPF-ranked SD rats were randomly divided into a control group, a model group, an EA group, an aconitine group and an EA plus aconitine group, with 6 rats in each group. The rat model of acute heart failure was established by infusion of high-dose propranolol hydrochloride solution into the right femoral vein. After stabilized for 10 min in the modeled rats, EA was exerted at "Neiguan" (PC 6), with disperse-dense wave, 2 Hz/15 Hz in frequency, 3 mA in intensity, for 30 min in the EA group and the EA plus aconitine group; aconitine solution (10 μg/kg) was injected from the left femoral veins in the rats in the aconitine group and the EA plus aconitine group. Hemodynamic indexes such as the left ventricular systolic pressure (LVSP) and the maximum rate of increase/decrease of left ventricular pressure (±dp/dt@*RESULTS@#Compared with the control group, LVSP and ±dp/dt@*CONCLUSION@#The intervention with electroacupuncture achieves the synergism/ attenuation effect of aconitine for the improvements in heart failure probably by up-regulating the expression of SERCA2a and down-regulating the expression of PLB in myocardial tissue.

Evaluation of the fibulin 5 gene polymorphism as a factor related to the occurrence of pelvic organ prolapse

SUMMARY OBJECTIVE Pelvic organ prolapse (POP) is a very frequent situation in our population that may lead to a significant decrease in patients' quality of life. Currently, we are looking for predictive factors for the development of POPs; thus, this study seeks to evaluate whether the Fibulin 5 polymorphism (FBLN5) is associated with the occurrence of POP. METHODS This is a cohort study with postmenopausal women who were divided into groups by POP stage: POP stages 0 and I (control group) and POP stages III and IV (case group). Subsequently, analyses of genetic polymorphisms of FBLN5 were performed using the Restriction Fragment Length Polymorphism (RFLP) technique. RESULTS A total of 292 women were included in the study. Pregnancy, parity and vaginal delivery in the patients, as well as in data described in the literature, were related to the occurrence of POP in the univariate analysis. However, after binary logistic regression, home birth and age remained independent risk factors for POP. We found no association between the FBLN5 polymorphism and the occurrence of POP (p = 0.371). CONCLUSION There was no association between the FBLN5 polymorphism and the occurrence of POP in Brazilian women.

RESUMO OBJETIVOS O prolapso de órgãos pélvicos (POP) é uma situação muito frequente em nossa população que pode levar a uma diminuição significativa da qualidade de vida dos pacientes. Atualmente, buscam-se fatores preditivos para o desenvolvimento de POPs e, assim, este estudo correlaciona um polimorfismo de Fibulina 5 (FBLN5) com a ocorrência da doença. MÉTODOS Estudo de coorte com mulheres na pós-menopausa, divididas por grupos pelos estádios 0 e I do POP (grupo controle) e POP III e IV (grupo caso). Posteriormente, análises do polimorfismo genético de FBLN5 foram realizadas utilizando a técnica de Polimorfismo de Comprimento de Fragmentos de Restrição (RFLP). RESULTADOS Um total de 292 mulheres foi incluído no estudo. Gestação, paridade e parto vaginal, como bem descritos na literatura, foram relacionados à ocorrência de POPs na análise univariada. No entanto, após a regressão logística binária, o parto domiciliar e a idade permaneceram como fatores de risco independentes para os POPs. Não encontramos associação deste polimorfismo FBLN5 com a ocorrência de POP (p=0,371). CONCLUSÃO Não houve associação deste polimorfismo FBLN5 com a ocorrência de POPs em mulheres brasileiras.

The effect of rosmarinic acid on deformities occurring in brain tissue by craniectomy method. Histopathological evaluation of IBA-1 and GFAP expressions

Abstract Purpose To investigate the role of Rosmarinic acid (RA) in the prevention of traumatic brain injury and the immunohistochemical analysis of IBA-1 and GFAP expressions. Methods Healthy male rats were randomly divided into 3 groups consisting of 10 rats. Groups were as follows; control group, traumatic brain injury (TBI) group, and TBI+RA group. After traumatic brain injury, blood samples were taken from the animals and analyzed with various biochemical markers. And then IBA-1 and GFAP expressions were evaluated immunohistochemically. Results Significant results were obtained in all biochemical parameters between groups. Immunohistochemical sections showed IBA-1 not only in microglia and macrophage activity but also in degenerative neurons in blood vessel endothelial cells. However, GFAP reaction and post-traumatic rosmarinic acid administration showed positive expression in astrocytes with regular structure around the blood vessel. Conclusion Rosmarinic acid in blood vessel endothelial cells showed that preserving the integrity of astrocytic structure in the blood brain barrier may be an important antioxidant.

Effect of periapical inflammation on calcium binding proteins and ERK in the trigeminal nucleus / Efecto de la inflamación periapical sobre las proteínas fijadoras de calcio y ERK en el núcleo trigeminal

Peripheral inflammation induces plastic changes in neurons and glia which are regulated by free calcium and calcium binding proteins (CaBP). One of the mechanisms associated with the regulation of intracellular calcium is linked to ERK (Extracellular Signal-Regulated Kinase) and its phosphorylated condition (pERK). ERK phosphorylation is important for intracellular signal transduction and participates in regulating neuroplasticity and inflammatory responses. The aim of this study is to analyse the expression of two CaBPs and pERK in astrocytes and neurons in rat trigeminal subnucleus caudalis (Vc) after experimental periapical inflammation on the left mandibular first molar. At seven days post-treatment, the periapical inflammatory stimulus induces an increase in pERK expression both in S100b positive astrocytes and Calbindin D28k positive neurons, in the ipsilateral Vc with respect to the contralateral side and control group. pERK was observed coexpressing with S100b in astrocytes and in fusiform Calbindin D28k neurons in lamina I. These results could indicate that neural plasticity and pain sensitization could be maintained by ERK activation in projection neurons at 7 days after the periapical inflammation.

La inflamación periférica induce cambios plásticos en las neuronas y en la glía, los cuales están regulados por el calcio libre y las proteínas fijadoras calcio (CaBP). Uno de los mecanismos asociados con la regulación del calcio intrace-lular está vinculado con la fosforilación de la pro teína quinasa ERK. Asimismo, ERK fosforilado es importante para la trans-ducción de señales intracelulares y participa en la regulación de la neuroplasticidad y las respuestas inflamatorias. El objetivo de este estudio es analizar la expresión de dos CaBPs y pERK en astrocitos y neuronas del subnúcleo caudal del trigémino (Vc) después de una inflamación periapical experimental en el primer molar inferior izquierdo en ratas. A los siete días posteriores al tratamiento, el estímulo inflamatorio periapical induce un aumento en la expresión de pERK, en el número de astrocitos positivos para la proteína marcadora astroglial S100b y en neuronas positivas para Calbindina D28k, en el Vc ipsilateral respecto del lado contralateral y el grupo de control. Además, se observó coexpresión de pERK tanto en astrocitos S100b positivos, como en neuronas fusiformes Calbindin D28k positivas, de la lámina I. Estas observaciones podrían indicar que la neuroplasticidad y la sensibilización al dolor podrían mantenerse mediante la activación de ERK en las neuronas de proyección a los 7 días de la inflamación periapical.

Pioneering studies on monogenic central precocious puberty

ABSTRACT Pubertal timing in humans is determined by complex interactions including hormonal, metabolic, environmental, ethnic, and genetic factors. Central precocious puberty (CPP) is defined as the premature reactivation of the hypothalamic-pituitary-gonadal axis, starting before the ages of 8 and 9 years in girls and boys, respectively; familial CPP is defined by the occurrence of CPP in two or more family members. Pioneering studies have evidenced the participation of genetic factors in pubertal timing, mainly identifying genetic causes of CPP in sporadic and familial cases. In this context, rare activating mutations were identified in genes of the kisspeptin excitatory pathway (KISS1R and KISS1 mutations). More recently, loss-of-function mutations in two imprinted genes (MKRN3 and DLK1) have been identified as important causes of familial CPP, describing novel players in the modulation of the hypothalamic-pituitary-gonadal axis in physiological and pathological conditions. MKRN3 mutations are the most common cause of familial CPP, and patients with MKRN3 mutations present clinical features indistinguishable from idiopathic CPP. Meanwhile, adult patients with DLK1 mutations present high frequency of metabolic alterations (overweight/obesity, early onset type 2 diabetes and hyperlipidemia), indicating that DLK1 may be a novel link between reproduction and metabolism. Arch Endocrinol Metab. 2019;63(4):438-44

Serum regucalcin is a useful indicator of liver injury severity in patients with hepatitis B virus-related liver diseases

Regucalcin is a soluble protein that is principally expressed in hepatocytes. Studies of regucalcin have mainly been conducted in animals due to a lack of commercially available kits. We aimed to develop an enzyme-linked immunosorbent assay (ELISA) to quantify serum regucalcin in patients with hepatitis B virus (HBV)-related disease. High-titer monoclonal antibodies and a polyclonal antibody to regucalcin were produced, a double-antibody sandwich ELISA method was established, and serum regucalcin was determined in 47 chronic hepatitis B (CHB) patients, 91 HBV-related acute-on-chronic liver failure (HBV-ACLF) patients, and 33 healthy controls. The ELISA demonstrated an appropriate linear range, and high levels of reproducibility, sensitivity, specificity, accuracy, and stability. The median serum regucalcin concentrations in HBV-ACLF and CHB patients were 5.46 and 3.76 ng/mL, respectively (P<0.01), which were much higher than in healthy controls (1.72 ng/mL, both P<0.01). For the differentiation of CHB patients and healthy controls, the area under curve (AUC) was 0.86 with a cut-off of 2.42 ng/mL, 85.7% sensitivity, and 78.8% specificity. In contrast, the AUC of alanine aminotransferase (ALT) was lower (AUC=0.80, P=0.01). To differentiate ACLF from CHB, the AUC was 0.72 with a cut-off of 4.26 ng/mL, 77.0% sensitivity, and 61.2% specificity while the AUC of ALT was 0.41 (P=0.07). Thus, we have developed an ELISA that is suitable for measuring serum regucalcin and have shown that serum regucalcin increased with the severity of liver injury due to HBV-related diseases, such that it appears to be more useful than ALT as a marker of liver injury.

Role of TGF-β1/ILK/FSP1 signaling pathway in cyclosporin A-induced epithelialmesenchymal transition in cultured renal tubular epithelial cells / 南方医科大学学报

OBJECTIVE@#To explore the role of transforming growth factor-β1/integrin-linked kinase/fibroblast-specific protein 1 (TGF- β1/ILK/FSP1) signaling pathway in cyclosporine A (CsA)-induced renal tubular epithelial cell transdifferentiation.@*METHODS@#Rat renal tubular epithelial NRK-52E cells were induced with 1 mg/L CsA, treated with TGF-β1 inhibitor (SB431542, 10 μmol/L), or transfected with the ILK-RNAi lentiviral expression vector (ILKshRNA) or a negative control vector before CsA induction. The expressions of TGF-β1, ILK and FSP-1 mRNAs and proteins in the cells were detected using real-time PCR and Western blotting. The positive cells for α-SMA expression were detected by immunohistochemistry.@*RESULTS@#Compared with the blank control cells, the cells treated with CsA showed significantly increased levels of TGF-β1, ILK and FSP-1 mRNAs and proteins ( < 0.05). The expressions of TGF-β1, ILK and FSP-1 were significantly lower in TGF-β1 inhibitor group than in CsA group ( < 0.05). The levels of ILK and FSP-1 were significantly decreased after shRNA-mediated ILK silencing ( < 0.05). The number of positive cells for -SMA was significantly lower in cells treated with SB431542 and in cells with ILK silencing than in the cells treated with CsA alone ( < 0.05).@*CONCLUSIONS@#The activation of TGF-β1/ILK/FSP-1 signaling pathway is an important mechanism for CsA-induced transdifferentiation in rat renal tubular epithelial cells. ILK participates in CsA-induced epithelialmesenchymal transition of renal tubular epithelial cells.

Expression of calponin-1 and its pathogenic role in systemic sclerosis / 南方医科大学学报

OBJECTIVE@#To investigate the expression of calponin-1 (CNN1) in systemic sclerosis (SSc) and its pathogenic role in fibrosis.@*METHODS@#Skin biopsy samples were collected from 19 patients with SSc and 21 healthy subjects. Real-time PCR was used to detect the expression of and mRNAs in the samples, and the protein expression of CNN1 was detected using immunohistochemistry. In cultured primary human dermal fibroblasts, expression was knocked down RNA interference, and the mRNA expression levels of and the fibrosis-related genes , , , , and were detected using real-time PCR; the proliferation of the cells was assessed using a real-time cell proliferation detection system.@*RESULTS@#Compared with that in samples from normal subjects, the expression of mRNA was significantly increased in the skin tissue of patients with SSc ( < 0.05) with a positive correlation with α-SMA (=0.7219, < 0.0001); the protein expression of CNN1 was also significantly increased in the skin tissue of patients with SSc. In cultured primary skin fibroblasts, the expression of CNN1 mRNA was positively correlated with and mRNA expressions (=0.6547, < 0.05; =0.6438, < 0.05). knockdown in the fibroblasts significantly inhibited the cell proliferation, obviously lowered the expressions of fibrosis-related genes, and reduced the protein expression of collagen.@*CONCLUSIONS@#The expression of is increased in the skin tissues of patients with SSc, and knockdown can reduce the activity of dermal fibroblasts, suggesting the close correlation of CNN1 with fibrosis in SSc.

Functional Analysis and Immunochemical Analyses of Ca²⁺ Homeostasis-Related Proteins Expression of Glaucoma-Induced Retinal Degeneration in Rats

The retinal degeneration resulting from elevated intraocular pressure was evaluated through functional and morphological analyses, for better understanding of the pathophysiology of glaucoma. Ocular hypertension was induced via unilateral episcleral venous cauterization in rats. Experimental time was set at 1 and 3 days, and 1, 2, 4, and 8 weeks post-operation. Retinal function was analyzed using electroretinography. For morphological analysis, retinal tissues were processed for immunochemistry by using antibodies against the calcium-sensing receptor and calcium-binding proteins. Apoptosis was analyzed using the TUNEL method and electron microscopy. Amplitudes of a- and b-wave in scotopic and photopic responses were found to be reduced in all glaucomatous retinas. Photopic negative response for ganglion cell function significantly reduced from 1-day and more significantly reduced in 2-week glaucoma. Calcium-sensing receptor immunoreactivity in ganglion cells remarkably reduced at 8 weeks; conversely, protein amounts increased significantly. Calcium-binding proteins immunoreactivity in amacrine cells clearly reduced at 8 weeks, despite of uneven changes in protein amounts. Apoptosis appeared in both photoreceptors and ganglion cells in 8-week glaucomatous retina. Apoptotic feature of photoreceptors was typical, whereas that of ganglion cells was necrotic in nature. These findings suggest that elevated intraocular pressure affects the sensitivity of photoreceptors and retinal ganglion cells, and leads to apoptotic death. The calcium-sensing receptor may be a useful detector for alteration of extracellular calcium levels surrounding the ganglion cells.

Early Activation of Astrocytes does not Affect Amyloid Plaque Load in an Animal Model of Alzheimer's Disease / 神经科学通报·英文版

Astrocytes are closely associated with Alzheimer's disease (AD). However, their precise roles in AD pathogenesis remain controversial. One of the reasons behind the different results reported by different groups might be that astrocytes were targeted at different stages of disease progression. In this study, by crossing hAPP (human amyloid precursor protein)-J20 mice with a line of GFAP-TK mice, we found that astrocytes were activated specifically at an early stage of AD before the occurrence of amyloid plaques, while microglia were not affected by this crossing. Activation of astrocytes at the age of 3-5 months did not affect the proteolytic processing of hAPP and amyloid plaque loads in the brains of hAPP-J20 mice. Our data suggest that early activation of astrocytes does not affect the deposition of amyloid β in an animal model of AD.

Cortical Inflammation is Increased in a DSS-Induced Colitis Mouse Model / 神经科学通报·英文版

While inflammatory bowel disease (IBD) might be a risk factor in the development of brain dysfunctions, the underlying mechanisms are largely unknown. Here, mice were treated with 5% dextran sodium sulfate (DSS) in drinking water and sacrificed on day 7. The serum level of IL-6 increased, accompanied by elevation of the IL-6 and TNF-α levels in cortical tissue. However, the endotoxin concentration in plasma and brain of mice with DSS-induced colitis showed a rising trend, but with no significant difference. We also found significant activation of microglial cells and reduction in occludin and claudin-5 expression in the brain tissue after DSS-induced colitis. These results suggested that DSS-induced colitis increases systemic inflammation which then results in cortical inflammation via up-regulation of serum cytokines. Here, we provide new information on the impact of colitis on the outcomes of cortical inflammation.

Effects of rosuvastatin in homocysteine induced mouse vascular smooth muscle cell dedifferentiation and endoplasmic reticulum stress and its mechanisms / 中国应用生理学杂志

OBJECTIVE@#To investigate the effect of rosuvastatin on homocysteine (Hcy) induced mousevascular smooth muscle cells(VSMCs) dedifferentiation and endoplasmic reticulum stress(ERS).@*METHODS@#VSMCs were co-cultured with Hcy and different concentration of rosuvastatin (0.1, 1.0 and 10 μmol/L). Cytoskeleton remodeling, VSMCs phenotype markers (smooth muscle actin-α, calponin and osteopontin) and ERS marker mRNAs (Herpud1, XBP1s and GRP78) were detected at predicted time. Tunicamycin was used to induce, respectively 4-phenylbutyrate(4-PBA) inhibition, ERS in VSMCs and cellular migration, proliferation and expression of phenotype proteins were analyzed. Mammalian target of rapamycin(mTOR)-P70S6 kinase (P70S6K) signaling agonist phosphatidic acid and inhibitor rapamycin were used in Rsv treated VSMCs. And then mTOR signaling and ERS associated mRNAs were detected.@*RESULTS@#Compared with Hcy group, Hcy+ Rsv group (1.0 and 10 μmol/L) showed enhanced α-SMA and calponin expression (<0.01), suppressed ERS mRNA levels (<0.01) and promoted polarity of cytoskeleton. Compared with Hcy group, Hcy+Rsv group and Hcy+4-PBA group showed suppressed proliferation, migration and enhanced contractile protein expression (<0.01); while tunicamycin could reverse the effect of Rsv on Hcy treated cells. Furthermore, alleviated mTOR-P70S6K phosphorylation and ERS (<0.01)were observed in Hcy+Rsv group and Hcy+rapamycin group, compared with Hcy group; while phosphatidic acid inhibited the effect of Rsv on mTOR signaling activation and ERS mRNA levels (<0.01).@*CONCLUSIONS@#Rosuvastatin could inhibit Hcy induced VSMCs dedifferentiation suppressing ERS, which might be regulated by mTOR-P70S6K signaling.