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1.
mBio ; 12(1)2021 01 05.
Artigo em Inglês | MEDLINE | ID: mdl-33402537

RESUMO

Invasive bacterial infections during pregnancy are a major risk factor for preterm birth, stillbirth, and fetal injury. Group B streptococci (GBS) are Gram-positive bacteria that asymptomatically colonize the lower genital tract but infect the amniotic fluid and induce preterm birth or stillbirth. Experimental models that closely emulate human pregnancy are pivotal for the development of successful strategies to prevent these adverse pregnancy outcomes. Using a unique nonhuman primate model that mimics human pregnancy and informs temporal events surrounding amniotic cavity invasion and preterm labor, we show that the animals inoculated with hyaluronidase (HylB)-expressing GBS consistently exhibited microbial invasion into the amniotic cavity, fetal bacteremia, and preterm labor. Although delayed cytokine responses were observed at the maternal-fetal interface, increased prostaglandin and matrix metalloproteinase levels in these animals likely mediated preterm labor. HylB-proficient GBS dampened reactive oxygen species production and exhibited increased resistance to neutrophils compared to an isogenic mutant. Together, these findings demonstrate how a bacterial enzyme promotes GBS amniotic cavity invasion and preterm labor in a model that closely resembles human pregnancy.IMPORTANCE Group B streptococci (GBS) are bacteria that commonly reside in the female lower genital tract as asymptomatic members of the microbiota. However, during pregnancy, GBS can infect tissues at the maternal-fetal interface, leading to preterm birth, stillbirth, or fetal injury. Understanding how GBS evade host defenses during pregnancy is key to developing improved preventive therapies for these adverse outcomes. In this study, we used a unique nonhuman primate model to show that an enzyme secreted by GBS, hyaluronidase (HylB) promotes bacterial invasion into the amniotic cavity and fetus. Although delayed immune responses were seen at the maternal-fetal interface, animals infected with hyaluronidase-expressing GBS exhibited premature cervical ripening and preterm labor. These observations reveal that HylB is a crucial GBS virulence factor that promotes bacterial invasion and preterm labor in a pregnancy model that closely emulates human pregnancy. Therefore, hyaluronidase inhibitors may be useful in therapeutic strategies against ascending GBS infection.


Assuntos
Hialuronoglucosaminidase/metabolismo , Neutrófilos/imunologia , Trabalho de Parto Prematuro/imunologia , Infecções Estreptocócicas/imunologia , Streptococcus agalactiae/metabolismo , Líquido Amniótico/microbiologia , Animais , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Hialuronoglucosaminidase/genética , Inflamação , Pulmão/microbiologia , Pulmão/patologia , Macaca nemestrina , Neutrófilos/microbiologia , Gravidez , Nascimento Prematuro , Primatas , Infecções Estreptocócicas/metabolismo , Infecções Estreptocócicas/microbiologia , Streptococcus agalactiae/enzimologia , Streptococcus agalactiae/genética , Streptococcus agalactiae/imunologia
2.
Medicine (Baltimore) ; 99(50): e23369, 2020 Dec 11.
Artigo em Inglês | MEDLINE | ID: mdl-33327261

RESUMO

Advanced studies demonstrated that hypoxic stress induced KIAA1199 expression leading to enhanced cell migration. KIAA1199 is a protein related with cancer metastasis. Hypoxia inducible factor 1α (HIF-1α) is a transcriptional factor that maintains oxygen homeostasis. Both KIAA1199 and HIF-1α were upregulated in many human cancers. In the present study, co-expression of KIAA1199 and HIF-1α was evaluated for the clinicopathological characteristics and survival in hepatocellular carcinoma (HCC). Clinical-pathological information and follow-up data were collected from 152 HCC patients. KIAA1199 and HIF-1α expression were scored based on the percentage and intensity of immunohistochemical staining in pathological slide. Correlations between clinical features and the expression of KIAA1199 and HIF-1α were evaluated by Chi-square test, Kaplan-Meier curves and multivariate Cox regression analysis. The frequency of KIAA1199 high expression was higher in HCC than adjacent tissue. KIAA1199(H)/HIF-1α(H) tumors were more frequently of TNM (P = .011), tumor size (P = .021), vascular invasion (P = .002) and HBV (P = .001). In survival analysis, KIAA1199(H)/HIF-1α(H) patients had the worst prognosis. Using the combination of the two parameters increased the prognostic value (P < .01 vs P = .03). KIAA1199 in combination with HIF-1α expression tends to indicate a more accurate prognosis.


Assuntos
Carcinoma Hepatocelular/genética , Regulação Neoplásica da Expressão Gênica/genética , Hialuronoglucosaminidase/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Neoplasias Hepáticas/genética , Adulto , Idoso , Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/mortalidade , Feminino , Humanos , Estimativa de Kaplan-Meier , Neoplasias Hepáticas/mortalidade , Masculino , Pessoa de Meia-Idade , Prognóstico , Modelos de Riscos Proporcionais
3.
J Cancer Res Clin Oncol ; 146(10): 2519-2534, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32648226

RESUMO

PURPOSE: Metastasis is an unavoidable event happened among almost all small cell lung cancer (SCLC) patients. However, the molecular driven factors have not been elucidated. Recently, a novel hydrolase called cell migration inducing hyaluronidase (CEMIP) triggered both migration and invasion in many tumors but not SCLC. Therefore, in this study, we verified that CEMIP promoted migration and invasion in SCLC and applied proteomics analysis to screen out potential target profiles and the signaling pathway related to CEMIP regulation. METHOD: Immunofluorescence was conducted to exam the expression of CEMIP on SCLC and paired adjacent normal tissues among enrollment. RT-qPCR and Western blot (WB) assays were conducted to valuate cellular protein and mRNA expression of CEMIP and EMT markers. Lentivirus-CEMIP-shRNAs and CEMIP plasmid were used for expression manipulating. Changes of cellular migration and invasion were tested through transwell assays. Tandem Mass Tag (TMT) peptide labeling coupled with LC-MS/MS was used for quantifying proteins affected by reducing expression of CEMIP on H446 cells. RESULTS: The expression of CEMIP showed 1.64 ± 0.16-fold higher in SCLC tissues than their normal counterpart. Decreasing the expression of CEMIP on SCLC cells H446 regressed both cellular migration and invasion ability, whereas the promoting cellular migration and invasion was investigated through over-expressing CEMIP on H1688. Proteomic and bioinformatics analysis revealed that total 215 differentially expressed proteins (DEPs) that either their increasing or decreasing relative expression met threshold of 1.2-fold changes with p value ≤ 0.05. The dramatic up-regulated DEPs included an unidentified peptide sequence (encoded by cDNA FLJ52096) SPICE1 and CRYAB, while the expression of S100A6 was largely down-regulated. DEPs mainly enriched on caveolae of cellular component, calcium ion binding of biological process and epithelial cell migration of molecular function. KEGG enrichment indicated that DEPs mainly exerted their function on TGF-ß, GABAergic synapse and MAPK signaling pathway. CONCLUSION: It is the first report illustrating that CEMIP might be one of the metastatic triggers in SCLC. And also, it provided possible molecular mechanism cue and potential downstream target on CEMIP-induced cellular migration and invasion on SCLC.


Assuntos
Hialuronoglucosaminidase/metabolismo , Neoplasias Pulmonares/metabolismo , Proteoma/metabolismo , Carcinoma de Pequenas Células do Pulmão/metabolismo , Idoso , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Feminino , Imunofluorescência , Humanos , Hialuronoglucosaminidase/biossíntese , Hialuronoglucosaminidase/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Estudos Retrospectivos , Transdução de Sinais , Carcinoma de Pequenas Células do Pulmão/genética , Carcinoma de Pequenas Células do Pulmão/patologia
4.
Hum Genet ; 139(11): 1471-1483, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32583022

RESUMO

Human growth is a complex trait determined by genetic factors in combination with external stimuli, including environment, nutrition and hormonal status. In the past, several genome-wide association studies (GWAS) have collectively identified hundreds of genetic variants having a putative effect on determining adult height in different worldwide populations. Theoretically, a valuable approach to better understand the mechanisms of complex traits as adult height is to study a population exhibiting extreme stature phenotypes, such as African Baka Pygmies. After phenotypic characterization, we sequenced the whole exomes of a cohort of Baka Pygmies and their non-Pygmies Bantu neighbors to highlight genetic variants associated with the reduced stature. Whole exome data analysis revealed 29 single nucleotide polymorphisms (SNPs) significantly associated with the reduced height in the Baka group. Among these variants, we focused on SNP rs7629425, located in the 5'-UTR of the Hyaluronidase-2 (HYAL2) gene. The frequency of the alternative allele was significantly increased compared to African and non-African populations. In vitro luciferase assay showed significant differences in transcription modulation by rs7629425 C/T alleles. In conclusion, our results suggested that the HYAL2 gene variants may play a role in the etiology of short stature in Baka Pygmies population.


Assuntos
Grupo com Ancestrais do Continente Africano/genética , Moléculas de Adesão Celular/genética , Proteínas Ligadas por GPI/genética , Transtornos do Crescimento/genética , Hialuronoglucosaminidase/genética , Polimorfismo de Nucleotídeo Único/genética , Adulto , Alelos , Estatura/genética , Exoma/genética , Feminino , Estudo de Associação Genômica Ampla/métodos , Humanos , Masculino
5.
PLoS One ; 15(3): e0230537, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32208452

RESUMO

During the blood feeding, sand fly females inject saliva containing immunomodulatory and anti-haemostatic molecules into their vertebrate hosts. The saliva composition is species-specific, likely due to an adaptation to particular haemostatic pathways of their preferred host. Research on sand fly saliva is limited to the representatives of two best-studied genera, Phlebotomus and Lutzomyia. Although the members of the genus Sergentomyia are highly abundant in many areas in the Old World, their role in human disease transmission remains uncertain. Most Sergentomyia spp. preferentially attack various species of reptiles, but feeding on warm-blooded vertebrates, including humans and domestic animals, has been repeatedly described, especially for Sergentomyia schwetzi, of which salivary gland transcriptome and proteome is analyzed in the current study. Illumina RNA sequencing and de novo assembly of the reads and their annotation revealed 17,293 sequences homologous to other arthropods' proteins. In the sialome, all proteins typical for sand fly saliva were identified-antigen 5-related, lufaxin, yellow-related, PpSP15-like, D7-related, ParSP25-like, and silk proteins, as well as less frequent salivary proteins included 71kDa-like, ParSP80-like, SP16-like, and ParSP17-like proteins. Salivary enzymes include apyrase, hyaluronidase, endonuclease, amylase, lipase A2, adenosine deaminase, pyrophosphatase, 5'nucleotidase, and ribonuclease. Proteomics analysis of salivary glands identified 631 proteins, 81 of which are likely secreted into the saliva. We also compared two S. schwetzi lineages derived from the same origin. These lineages were adapted for over 40 generations for blood feeding either on mice (S-M) or geckos (S-G), two vertebrate hosts with different haemostatic mechanisms. Altogether, 20 and 40 annotated salivary transcripts were up-regulated in the S-M and S-G lineage, respectively. Proteomic comparison revealed ten salivary proteins more abundant in the lineage S-M, whereas 66 salivary proteins were enriched in the lineage S-G. No difference between lineages was found for apyrase activity; contrarily the hyaluronidase activity was significantly higher in the lineage feeding on mice.


Assuntos
Proteínas de Insetos/genética , Psychodidae/genética , Glândulas Salivares/metabolismo , Transcriptoma , Animais , Apirase/análise , Apirase/genética , Apirase/metabolismo , Hialuronoglucosaminidase/análise , Hialuronoglucosaminidase/genética , Hialuronoglucosaminidase/metabolismo , Proteínas de Insetos/análise , Proteínas de Insetos/metabolismo , Lagartos , Camundongos , Filogenia , Psychodidae/metabolismo , Receptores Odorantes/análise , Receptores Odorantes/genética , Receptores Odorantes/metabolismo
6.
Appl Microbiol Biotechnol ; 104(4): 1621-1632, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31907577

RESUMO

Hyaluronidases that break down hyaluronan are widely used for preparation of low molecular weight hyaluronan. Leech hyaluronidase (LHyal) is a newly discovered hyaluronidase with outstanding enzymatic properties. The Pichia pastoris expression system of LHyal that depends on AOX1 promoter (PAOX1) has been constructed. However, the addition of the toxic inducer methanol is a big safety concern. Here, a combinational strategy was adopted for constitutive expression of LHyal to high level in P. pastoris. By optimizing the combination of promoters PGAP, PGAP(m), and PTEF1 and signal peptides α-factor, nsB, and sp23, the enzyme activity of extracellular LHyal reached 1.38 × 105 U/mL in shake flasks. N-terminal engineering with neutral polar amino acids further increased LHyal activity to 2.06 × 105 U/mL. In addition, the impact of overexpressing transcription factors Aft1, Gal4-like, and Yap1 on LHyal production was also investigated. We found the co-expression of Aft1 significantly enhanced the expression of LHyal to 3.03 × 105 U/mL. Finally, LHyal activity of 2.12 × 106 U/mL was achieved in a 3-L fermenter, with a high productivity of 1.96 × 104 U/mL/h. The engineered LHyal-producing Pichia pastoris strains will be more attractive for production of hyaluronidase on industrial scale.


Assuntos
Hialuronoglucosaminidase/biossíntese , Sanguessugas/enzimologia , Pichia/metabolismo , Animais , Técnicas de Cultura Celular por Lotes , Reatores Biológicos , Hialuronoglucosaminidase/genética , Microbiologia Industrial , Sanguessugas/genética , Pichia/genética , Regiões Promotoras Genéticas , Sinais Direcionadores de Proteínas/genética , Fatores de Transcrição/genética
7.
Genes Chromosomes Cancer ; 59(5): 309-317, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-31898851

RESUMO

Myxoinflammatory fibroblastic sarcoma (MIFS) has recurrent genetic features in the form of a translocation t(1;10)(p22-31;q24-25), BRAF gene fusions, and/or an amplicon in 3p11-12 including the VGLL3 gene. The breakpoints on chromosomes 1 and 10 in the t(1;10) cluster in or near the TGFBR3 and OGA genes, respectively. We here used a combination of deep sequencing of the genome (WGS), captured sequences (Cap-seq), and transcriptome (RNA-seq) and genomic arrays to investigate the molecular outcome of the t(1;10) and the VGLL3 amplicon, as well as to assess the spectrum of other recurrent genomic features in MIFS. Apart from a ROBO1-BRAF chimera in a t(1;10)-negative MIFS-like tumor, no fusion gene was found at RNA-seq. This was in line with WGS and Cap-seq results, revealing variable breakpoints in chromosomes 1 and 10 and genomic breakpoints that should not yield functional fusion transcripts. The most common genomic rearrangements were breakpoints in or around the OGA, NPM3, and FGF8 genes in chromosome band 10q24, and loss of 1p11-p21 and 10q26-qter (all simultaneously present in 6/7 MIFS); a breakpoint in or near TGFBR3 in chromosome 1 was found in four of these tumors. Amplification and overexpression of VGLL3 was a consistent feature in MIFS and MIFS-like tumors with amplicons in 3p11-12. The significant molecular genetic outcome of the recurrent t(1;10) could be loss of genetic material from 1p and 10q. Other recurrent genomic imbalances in MIFS, such as homozygous loss of CDKN2A and 3p- and 13q-deletions, are shared with other sarcomas, suggesting overlapping pathogenetic pathways.


Assuntos
Biomarcadores Tumorais/genética , Fibrossarcoma/genética , Mixossarcoma/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos de Neoplasias/genética , Cromossomos Humanos Par 1 , Cromossomos Humanos Par 10 , Feminino , Fibrossarcoma/patologia , Rearranjo Gênico , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Histona Acetiltransferases/genética , Humanos , Hialuronoglucosaminidase/genética , Masculino , Pessoa de Meia-Idade , Mixossarcoma/patologia , Receptores de Fatores de Crescimento Transformadores beta/genética , Fatores de Transcrição/genética , Translocação Genética
8.
J Biol Chem ; 295(8): 2483-2494, 2020 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-31949043

RESUMO

The immune-regulatory compound histamine is involved in the metabolism of the essential skin component hyaluronan (HA). We previously reported that histamine up-regulates the expression of HYBID (hyaluronan-binding protein involved in hyaluronan depolymerization, also called CEMIP or KIAA1199), which plays a key role in HA degradation. However, no information is available about histamine's effects on HA synthase (HAS) expression, the molecular sizes of HA species produced, and histamine receptors and their signaling pathways in skin fibroblasts. Moreover, histamine's effects on photoaged skin remain elusive. Here, we show that histamine increases HA degradation by up-regulating HYBID and down-regulating HAS2 in human skin fibroblasts in a dose- and time-dependent manner and thereby decreases the total amounts and sizes of newly produced HA. Histamine H1 blocker abrogated the histamine effects on HYBID up-regulation, HAS2 suppression, and HA degradation. Histamine H1 agonist exhibited effects on HA levels, composition, and breakdown similar to those of histamine. Of note, blockade of protein kinase Cδ or PI3K-Akt signaling abolished histamine-mediated HYBID stimulation and HAS2 suppression, respectively. Immunohistochemical experiments revealed a significant ∼2-fold increase in tryptase-positive mast cells in photoaged skin, where HYBID and HAS2 expression levels were increased and decreased, respectively, compared with photoprotected skin. These results indicate that histamine controls HA metabolism by up-regulating HYBID and down-regulating HAS2 via distinct signaling pathways downstream of histamine receptor H1. They further suggest that histamine may contribute to photoaged skin damage by skewing HA metabolism toward degradation.


Assuntos
Fibroblastos/metabolismo , Histamina/farmacologia , Hialuronan Sintases/metabolismo , Ácido Hialurônico/metabolismo , Hialuronoglucosaminidase/metabolismo , Pele/citologia , Linhagem Celular , Fibroblastos/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Hialuronan Sintases/genética , Hialuronoglucosaminidase/genética , Mastócitos/efeitos dos fármacos , Mastócitos/metabolismo , Peso Molecular , Fosfatidilinositol 3-Quinases/metabolismo , Proteína Quinase C-delta/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Histamínicos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Envelhecimento da Pele/efeitos dos fármacos , Fatores de Tempo
9.
Sci Rep ; 10(1): 280, 2020 01 14.
Artigo em Inglês | MEDLINE | ID: mdl-31937874

RESUMO

Chronic hypoxia leads to pathologic remodeling of the pulmonary vasculature and pulmonary hypertension (PH). The antioxidant enzyme extracellular superoxide dismutase (SOD3) protects against hypoxia-induced PH. Hyaluronan (HA), a ubiquitous glycosaminoglycan of the lung extracellular matrix, is rapidly recycled at sites of vessel injury and repair. We investigated the hypothesis that SOD3 preserves HA homeostasis by inhibiting oxidative and enzymatic hyaluronidase-mediated HA breakdown. In SOD3-deficient mice, hypoxia increased lung hyaluronidase expression and activity, hyaluronan fragmentation, and effacement of HA from the vessel wall of small pulmonary arteries. Hyaluronan fragmentation corresponded to hypoxic induction of the cell surface hyaluronidase-2 (Hyal2), which was localized in the vascular media. Human pulmonary artery smooth muscle cells (HPASMCs) demonstrated hypoxic induction of Hyal2 and SOD-suppressible hyaluronidase activity, congruent to our observations in vivo. Fragmentation of homeostatic high molecular weight HA promoted HPASMC proliferation in vitro, whereas pharmacologic inhibition of hyaluronidase activity prevented hypoxia- and oxidant-induced proliferation. Hypoxia initiates SOD3-dependent alterations in the structure and regulation of hyaluronan in the pulmonary vascular extracellular matrix. These changes occurred soon after hypoxia exposure, prior to appearance of PH, and may contribute to the early pathogenesis of this disease.


Assuntos
Ácido Hialurônico/metabolismo , Hipertensão Pulmonar/patologia , Hipóxia , Animais , Hipóxia Celular , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Ácido Hialurônico/análise , Ácido Hialurônico/farmacologia , Hialuronoglucosaminidase/antagonistas & inibidores , Hialuronoglucosaminidase/genética , Hialuronoglucosaminidase/metabolismo , Hipertensão Pulmonar/metabolismo , Pulmão/enzimologia , Pulmão/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/metabolismo , Artéria Pulmonar/citologia , Artéria Pulmonar/enzimologia , Superóxido Dismutase/deficiência , Superóxido Dismutase/genética , Regulação para Cima
10.
Biomed Pharmacother ; 123: 109728, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31846842

RESUMO

BACKGROUND: H3K27me3 modification inactivates gene transcription by resulting in condensed chromatin. However, the landscape and biological functions of H3K27me3 in breast cancer remain unclear. METHODS: Fluorescence enzyme assay was used to analyze the cell proliferation. Transwell assay was used to test the ability of migration and invasion in MDA-MB-231 cells with designed treatment. Transfection of exogenous plasmid was used to intervene specific gene expression. Nude mouse tumor xenograft model was employed to detect the effect of GSKJ-4 in vivo. ChIP-Seq analyzed the modification state of H3K27me3 around the TSS of the gene CEMIP. RNA-Seq was used to analyze the mRNA levels after treating with GSKJ-4 in MDA-MB-231 cells. RESULTS: Loss of H3K27me3 is specific for aggressive subtypes of breast cancer and may be a useful diagnostic marker. Epigenetic chemical screening identified histone H3K27me3 demethylation inhibition as a therapeutic strategy for triple-negative breast cancer (TNBC). Functional studies and RNA-seq/ChIP-seq data revealed that inactivation of the protein CEMIP (which is translated by oncogene KIAA1199) by increasing H3K27me3 leads to decreased tumor cell growth and migration. Moreover, survival analysis showed that CEMIP was associated with poor outcome in TNBC. CONCLUSIONS: Our data suggest H3K27me3 loss as an important event in CEMIP mediated breast cancer carcinogenesis and progression. Loss of H3K27me3 is specific for aggressive subtypes of breast cancer and may be a useful diagnostic marker.


Assuntos
Benzazepinas/farmacologia , Neoplasias da Mama/patologia , Proliferação de Células/efeitos dos fármacos , Histonas/metabolismo , Hialuronoglucosaminidase/metabolismo , Pirimidinas/farmacologia , Animais , Biomarcadores Tumorais , Neoplasias da Mama/tratamento farmacológico , Carcinogênese , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Estudo de Associação Genômica Ampla , Histonas/genética , Humanos , Hialuronoglucosaminidase/genética , Camundongos , Camundongos Nus , Prognóstico , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
11.
Oncol Rep ; 43(1): 328-336, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31746434

RESUMO

Aberrant expression of long noncoding RNAs (lncRNAs) has been demonstrated in human cancers and regulates the malignant behavior of cancer cells. Previous studies demonstrated the critical involvement of lncRNA histocompatibility leukocyte antigen (HLA) complex P5 (HCP5) in the development of cancers, however, the function of HCP5 in prostate cancer has not been reported. In the present study, we found the overexpressed expression of HCP5 in prostate cancer tissues and cell lines via RT­qPCR analysis. High expression of HCP5 was positively correlated with the metastasis of prostate cancer. Downregulation of HCP5 inhibited the proliferation, colony formation and induced apoptosis of prostate cancer cells. Functional experiments demonstrated that HCP5 acted as a competing endogenous RNA (ceRNA) to sponge miR­4656. Ectopic expression of HCP5 decreased the expression of miR­4656 in prostate cancer cells. MiR­4656 was found to be decreased in prostate cancer tissues and was negatively correlated with the expression of HCP5. Further luciferase reporter assay revealed that miR­4656 was able to bind the 3'­untranslated region (3'­UTR) of the cell migration inducing hyaluronidase 1 (CEMIP) and suppressed the expression of CEMIP. Consistent with the negative regulation of miR­4656 by HCP5, western blot analysis uncovered that overexpression of HCP5 upregulated the abundance of CEMIP in prostate cancer cells. The CCK­8 assay showed that depletion of CEMIP significantly inhibited the HCP5­promoted proliferation of prostate cancer cells. Collectively, our data provide a novel mechanism by which HCP5 regulates the progression of prostate cancer.


Assuntos
Hialuronoglucosaminidase/genética , MicroRNAs/genética , Neoplasias da Próstata/patologia , RNA Longo não Codificante/genética , Regiões 3' não Traduzidas , Idoso , Animais , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Metástase Neoplásica , Transplante de Neoplasias , Células PC-3 , Neoplasias da Próstata/genética , Regulação para Cima
12.
Anticancer Res ; 39(12): 6567-6573, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31810922

RESUMO

BACKGROUND/AIM: The KIAA1199 gene has been associated with cancer-cell proliferation, but its functions remain poorly studied. Here, we examined the clinical significance of the KIAA1199 mRNA levels in locally advanced gastric cancer (GC). Materials and Methods/Results: Using samples from 254 patients with stage II/III GC, we found significantly higher KIAA1199 levels in cancerous tissues compared to adjacent normal mucosa (ANM). There was no significant relationship between KIAA1199 expression and clinical features. Although overall survival rates (OSR) of patients, who underwent surgery did not correlate with KIAA1199 expression, patients who underwent adjuvant chemotherapy with S-1 and had high KIAA1199 levels displayed significantly lower OSR. KIAA1199 knock down (KIAA1199-KD) suppressed proliferation, invasiveness, and sensitivity of GC cells to 5-fluorouracil (5-FU). CONCLUSION: KIAA1199 expression appears to be a promising prognostic marker in patients with locally advanced GC, who underwent postoperative adjuvant chemotherapy with S-1. KIAA1199 may represent a novel target for GC pharmacotherapy.


Assuntos
Hialuronoglucosaminidase/genética , Ácido Oxônico/uso terapêutico , Neoplasias Gástricas/terapia , Tegafur/uso terapêutico , Regulação para Cima , Idoso , Linhagem Celular Tumoral , Proliferação de Células , Quimioterapia Adjuvante , Procedimentos Cirúrgicos do Sistema Digestório , Combinação de Medicamentos , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Prognóstico , Neoplasias Gástricas/genética , Neoplasias Gástricas/mortalidade , Análise de Sobrevida
13.
Vet J ; 254: 105393, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31836163

RESUMO

Mammary gland tumors are a heterogeneous group of neoplastic diseases. Genetic studies make it possible to determine genetic profiles and identify new molecular markers. The aim of the study was to evaluate the gene expression profile of canine mammary carcinomas and identify potential prognostic markers. Twelve mammary cancer samples from bitches were collected for the evaluation of global gene expression. Microarray assays were performed using commercial kits. Statistical analysis of the microarray was done using moderate t-statistic and adjusted using the Benjamini and Hochberg procedure. Differential connectivity analysis was also performed. Enrichment analyses were conducted using WebGestalt. P-values were calculated using hypergeometric statistics and adjusted using the Benjamini and Hochberg procedure. The HYAL-1 gene was validated using quantitative PCR (qPCR). There were 878 upregulated genes and 821 downregulated genes in the neoplasms studied. Enrichment analysis (individual analysis) identified the HYAL-1 gene as a potential marker of tumorigenesis and tumor recurrence. Differential connectivity analysis demonstrated 262 differentially connected genes.


Assuntos
Doenças do Cão/genética , Neoplasias Mamárias Animais/genética , Animais , Biomarcadores Tumorais/metabolismo , DNA de Neoplasias , Doenças do Cão/enzimologia , Cães , Feminino , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Hialuronoglucosaminidase/genética , Hialuronoglucosaminidase/metabolismo , Neoplasias Mamárias Animais/enzimologia , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase em Tempo Real/veterinária
14.
FASEB J ; 33(12): 14440-14449, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31670981

RESUMO

The glycosylphosphatidylinositol-anchored sperm hyaluronidases (Hyals), sperm adhesion molecule 1 (SPAM1) and HYAL5, have long been believed to assist in sperm penetration through the cumulus-oocyte complex (COC), but their role in mammalian fertilization remains unclear. Previously, we have shown that mouse sperm devoid of either Spam1 or Hyal5 are still capable of penetrating the COC and that the loss of either Spam1 or Hyal5 alone does not cause male infertility in mice. In the present study, we found that Spam1/Hyal5 double knockout (dKO) mice produced significantly fewer offspring compared with wild-type (WT) mice, and this was due to defective COC dispersal. A comparative analysis between WT and Spam1/Hyal5 dKO epididymal sperm revealed that the absence of these 2 sperm Hyals resulted in a marked accumulation of sperm on the outside of the COC. This impaired sperm activity is likely due to the deficiency in the sperm Hyals, even though other somatic Hyals are expressed normally in the dKO mice. The fertilization ability of the Spam1/Hyal5 dKO sperm was restored by adding purified human sperm Hyal to the in vitro fertilization medium. Our results suggest that Hyal deficiency in sperm may be a significant risk factor for male sterility.-Park, S., Kim, Y.-H., Jeong, P.-S., Park, C., Lee, J.-W., Kim, J.-S., Wee, G., Song, B.-S., Park, B.-J., Kim, S.-H., Sim, B.-W., Kim, S.-U., Triggs-Raine, B., Baba, T., Lee, S.-R., Kim, E. SPAM1/HYAL5 double deficiency in male mice leads to severe male subfertility caused by a cumulus-oocyte complex penetration defect.


Assuntos
Moléculas de Adesão Celular/metabolismo , Hialuronoglucosaminidase/metabolismo , Infertilidade Masculina/genética , Interações Espermatozoide-Óvulo/genética , Interações Espermatozoide-Óvulo/fisiologia , Espermatozoides/fisiologia , Animais , Moléculas de Adesão Celular/genética , Células do Cúmulo , Hialuronoglucosaminidase/genética , Masculino , Camundongos , Camundongos Knockout , Oócitos
15.
Nat Cell Biol ; 21(11): 1403-1412, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31685984

RESUMO

The development of effective therapies against brain metastasis is currently hindered by limitations in our understanding of the molecular mechanisms driving it. Here we define the contributions of tumour-secreted exosomes to brain metastatic colonization and demonstrate that pre-conditioning the brain microenvironment with exosomes from brain metastatic cells enhances cancer cell outgrowth. Proteomic analysis identified cell migration-inducing and hyaluronan-binding protein (CEMIP) as elevated in exosomes from brain metastatic but not lung or bone metastatic cells. CEMIP depletion in tumour cells impaired brain metastasis, disrupting invasion and tumour cell association with the brain vasculature, phenotypes rescued by pre-conditioning the brain microenvironment with CEMIP+ exosomes. Moreover, uptake of CEMIP+ exosomes by brain endothelial and microglial cells induced endothelial cell branching and inflammation in the perivascular niche by upregulating the pro-inflammatory cytokines encoded by Ptgs2, Tnf and Ccl/Cxcl, known to promote brain vascular remodelling and metastasis. CEMIP was elevated in tumour tissues and exosomes from patients with brain metastasis and predicted brain metastasis progression and patient survival. Collectively, our findings suggest that targeting exosomal CEMIP could constitute a future avenue for the prevention and treatment of brain metastasis.


Assuntos
Neoplasias Encefálicas/genética , Exossomos/metabolismo , Regulação Neoplásica da Expressão Gênica , Hialuronoglucosaminidase/genética , Neovascularização Patológica/genética , Microambiente Tumoral/genética , Animais , Encéfalo/metabolismo , Encéfalo/patologia , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/mortalidade , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Quimiocina CCL1/genética , Quimiocina CCL1/metabolismo , Quimiocina CXCL1/genética , Quimiocina CXCL1/metabolismo , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Exossomos/patologia , Humanos , Hialuronoglucosaminidase/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Metástase Neoplásica , Neovascularização Patológica/metabolismo , Neovascularização Patológica/mortalidade , Neovascularização Patológica/patologia , Transdução de Sinais , Análise de Sobrevida , Carga Tumoral , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Med Sci Monit ; 25: 6788-6796, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31501407

RESUMO

BACKGROUND KIAA1199 has been reported to be associated with malignant progression and poor clinical outcomes in various human malignancies. However, its clinical role and molecular function remain unknown in papillary thyroid cancer (PTC). MATERIAL AND METHODS The Cancer Genome Atlas (TCGA) was used to investigate the expression profiles of KIAA1199 and miR-486-5p in PTC. Immunohistochemistry was used to validate the protein expression of KIAA1199 in PTC. The Weighted Gene Co-expression Network Analysis (WGCNA) and Gene Set Enrichment Analysis (GSEA) were used to explore the potential pathway underling KIAA1199 in PTC. In vitro and in vivo experiments were performed to investigate the biological role of KIAA1199 in PTC progression. Luciferase reporter assays and Western blot analysis were performed to determine whether KIAA1199 is a downstream target of miR-486-5p. RESULTS We found that KIAA1199 was aberrantly elevated in PTC tissues compared with normal tissues, and upregulation of KIAA1199 was positively correlated with more advanced clinical variables. Additionally, bioinformatic analysis indicated that KIAA1199 was involved in cell migration and invasion. KIAA1199 silencing inhibited the invasive ability of PTC cells by affecting epithelial-mesenchymal transition (EMT) in vitro and in vivo. Furthermore, miR-486-5p was identified as an upstream microRNA that directly targets the 3'-UTR region of KIAA1199. CONCLUSIONS The miR-486-5p/KIAA1199/EMT axis might play a critical role in PTC invasion and metastasis and offers a potential therapeutic strategy for PTC.


Assuntos
Transição Epitelial-Mesenquimal/genética , Hialuronoglucosaminidase/metabolismo , MicroRNAs/metabolismo , Câncer Papilífero da Tireoide/genética , Câncer Papilífero da Tireoide/patologia , Animais , Sequência de Bases , Linhagem Celular Tumoral , Movimento Celular/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Hialuronoglucosaminidase/genética , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , Invasividade Neoplásica , Regulação para Cima/genética
17.
Oncol Rep ; 42(5): 2065-2074, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31545463

RESUMO

O­linked ß­N­acetylglucosamine (O­GlcNAc) modification is a dynamic post­translational modification process that is involved in many crucial biological processes, including cell cycle regulation, nutrient metabolism and extracellular signaling. This dynamic modification is dependent on the ambient glucose concentration and is catalyzed and removed by O­GlcNAc transferase (OGT) and O­GlcNAcase (OGA), respectively. The present study aimed to determine the role of O­GlcNAcylation during embryo implantation by inhibiting or enhancing its function and expression. The results revealed that the expression of O­GlcNAc­modified proteins in the human secretory endometrium was higher than that of the endometrium during the proliferative phase, as determined via western blotting and immunohistochemistry. Additionally, the level of endometrial O­GlcNAc modification increased gradually from the pre­receptive to the receptive phase, which was then decreased during the non­receptive phase. In endometrial cells, RNA interference was utilized to reduce the expression of two key O­GlcNAc synthesis and decomposition enzymes, OGT and OGA, to indirectly increase or decrease levels of O­GlcNAc modification. The results revealed that increasing the level of O­GlcNAc modification enhanced cellular proliferation, migration, invasion and adhesion, thereby promoting embryo implantation. It is hypothesized that O­GlcNAc modification serves an important role in the regulation of endometrial receptivity and embryo implantation. The results of the present study may have important implications for the understanding of female fertility and may help improve infertility treatments.


Assuntos
Acetilglucosamina/metabolismo , Antígenos de Neoplasias/metabolismo , Endométrio/metabolismo , Histona Acetiltransferases/metabolismo , Hialuronoglucosaminidase/metabolismo , N-Acetilglucosaminiltransferases/metabolismo , Adulto , Antígenos de Neoplasias/genética , Linhagem Celular , Movimento Celular , Proliferação de Células , Implantação do Embrião , Feminino , Fase Folicular/metabolismo , Glicosilação , Histona Acetiltransferases/genética , Humanos , Hialuronoglucosaminidase/genética , Pessoa de Meia-Idade , N-Acetilglucosaminiltransferases/genética
18.
J Microbiol Biotechnol ; 29(8): 1310-1315, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31370115

RESUMO

Hyaluronidases enhance therapeutic drug transport by breaking down the hyaluronan barrier to lymphatic and capillary vessels, facilitating their tissue absorption. Commercially available hyaluronidases are bovine in origin; however, they pose risks such as bovine spongiform encephalopathy. The present study aimed to develop a novel, highly active hyaluronidase and assess its function. Therefore, in order to find the most efficient active hyaluronidase, we produced several shortened hyaluronidases with partial removal of the N- or C-terminal regions. Moreover, we created an enzyme that connected six histidines onto the end of the hyaluronidase C-terminus. This simplified subsequent purification using Ni2+ affinity chromatography, making it feasible to industrialize this highly active recombinant hyaluronidase which exhibited catalytic activity equal to that of the commercial enzyme. Therefore, this simple and effective isolation method could increase the availability of recombinant hyaluronidase for research and clinical purposes.


Assuntos
Histidina/metabolismo , Hialuronoglucosaminidase/genética , Hialuronoglucosaminidase/metabolismo , Oligopeptídeos/metabolismo , Proteínas Recombinantes , Animais , Bovinos , Moléculas de Adesão Celular/metabolismo , Clonagem Molecular , Estabilidade Enzimática , Células HEK293 , Humanos , Ácido Hialurônico/metabolismo , Hialuronoglucosaminidase/isolamento & purificação , Concentração de Íons de Hidrogênio , Temperatura
19.
PLoS One ; 14(7): e0218736, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31260471

RESUMO

LL-37 is the only human cathelicidin-family host defense peptide and has been reported to interact with invading pathogens causing inflammation at various body sites. Recent studies showed high levels of LL-37 in the synovial-lining membrane of patients with rheumatoid arthritis, a common type of inflammatory arthritis. The present study aims to investigate the role of LL-37 on mechanisms associated with pathogenesis of inflammatory arthritis. The effects of LL-37 on the expression of proinflammatory cytokines, hyaluronan (HA) metabolism-related genes, cell death-related pathways, and cell invasion were investigated in SW982, a human synovial sarcoma cell line. Time-course measurements of proinflammatory cytokines and mediators showed that LL-37 significantly induced IL6 and IL17A mRNA levels at early time points (3-6 hr). HA-metabolism-related genes (i.e., HA synthase 2 (HAS2), HAS3, hyaluronidase 1 (HYAL1), HYAL2, and CD44) were co-expressed in parallel. In combination, LL-37 and IL17A significantly enhanced PTGS2, TNF, and HAS3 gene expression concomitantly with the elevation of their respective products, PGE2, TNF, and HA. Cell invasion rates and FN1 gene expression were also significantly enhanced. However, LL-37 alone or combined with IL17A did not affect cell mortality or cell cycle. Treatment of SW982 cells with both LL-37 and IL17A significantly enhanced IKK and p65 phosphorylation. These findings suggest that the chronic production of a high level of LL-37 may synchronize with its downstream proinflammatory cytokines, especially IL17A, contributing to the co-operative enhancement of pathogenesis mechanisms of inflammatory arthritis, such as high production of proinflammatory cytokines and mediators together with the activation of HA-metabolism-associated genes and cell invasion.


Assuntos
Peptídeos Catiônicos Antimicrobianos/farmacologia , Fibroblastos/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Ácido Hialurônico/metabolismo , Interleucina-17/farmacologia , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/imunologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/imunologia , Combinação de Medicamentos , Sinergismo Farmacológico , Fibroblastos/imunologia , Fibroblastos/patologia , Fibronectinas/genética , Fibronectinas/imunologia , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/imunologia , Regulação da Expressão Gênica/imunologia , Humanos , Receptores de Hialuronatos/genética , Receptores de Hialuronatos/imunologia , Hialuronan Sintases/genética , Hialuronan Sintases/imunologia , Ácido Hialurônico/imunologia , Hialuronoglucosaminidase/genética , Hialuronoglucosaminidase/imunologia , Quinase I-kappa B/genética , Quinase I-kappa B/imunologia , Inflamação , Interleucina-6/genética , Interleucina-6/imunologia , Transdução de Sinais , Membrana Sinovial/imunologia , Membrana Sinovial/patologia , Fator de Transcrição RelA/genética , Fator de Transcrição RelA/imunologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia
20.
Int Immunopharmacol ; 73: 203-211, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31103876

RESUMO

The overproduction of proteolytic enzymes and dysregulation of extracellular matrix (ECM) metabolism have been shown to accelerate the degradation process of articular cartilage. The purpose of this study was to investigate the role of KIAA1199 and its association with the pathophysiology of osteoarthritis (OA). We found that the expression of KIAA1199 was significantly upregulated in OA cartilage compared with normal tissues. Serum levels of KIAA1199 were higher in OA patients than in non-OA patients. Furthermore, knockdown of KIAA1199 inhibited interleukin-1 beta (IL-1ß)-induced ECM metabolic imbalance by regulating the expression of A disintegrin-like and metalloprotease (reprolysin type) with thrombospondin type 1 motif, 5; matrix metallopeptidase-13; aggrecan; and COL2A1. In addition, silencing of KIAA1199 significantly decreased the expression of inflammatory mediators such as prostaglandin E2, IL-6, and TNF-α. Mechanistic analyses further revealed that IL-1ß-induced activation of the Wnt/ß-catenin pathway was suppressed during KIAA1199 knockdown. Moreover, KIAA1199 expression was also upregulated in an in vivo rat OA model. Together, these results increase our understanding of the emerging role of KIAA1199 in the process of OA degeneration, and may lead to a novel molecular target to prevent cartilage degradation.


Assuntos
Cartilagem Articular/metabolismo , Condrócitos/metabolismo , Hialuronoglucosaminidase/metabolismo , Interleucina-1beta , Osteoartrite/metabolismo , Via de Sinalização Wnt , Animais , Células Cultivadas , Técnicas de Silenciamento de Genes , Humanos , Hialuronoglucosaminidase/sangue , Hialuronoglucosaminidase/genética , Masculino , Ratos Sprague-Dawley , Regulação para Cima
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