RESUMO
The objective of this study is to discern the proteomic differences responsible for hampering the receptivity of endometrium and subduing the fertility of females with polycystic ovary syndrome in analogy to healthy fertile females. This study was designed in collaboration with Hakeem Abdul Hameed Centenary Hospital affiliated to Jamia Hamdard, New Delhi, India. Serum samples were taken from infertile PCOS subjects (n = 6) and fertile control subjects (n = 6) whereas endometrial tissue samples were recruited from ovulatory PCOS (n = 4), anovulatory PCOS (n = 4) and normal healthy fertile control subjects (n = 4) for proteomic studies. Additionally, endometrial biopsies from healthy fertile control (n = 8), PCOS with infertility (n = 6), unexplained infertility (n = 3) and endometrial hyperplasia (n = 3) were taken for validation studies. Anthropometric, biochemical and hormonal evaluation was done for all the subjects enrolled in this study. Protein profiles were generated through 2D-PAGE and differential proteins analyzed with PD-QUEST software followed by identification with MALDI-TOF MS protein mass fingerprinting. Validation of identified proteins was done through RT-PCR relative expression analysis. Protein profiling of serum revealed differential expression of proteins involved in transcriptional regulation, embryogenesis, DNA repair, decidual cell ploidy, immunomodulation, intracellular trafficking and degradation processes. Proteins involved in cell cycle regulation, cellular transport and signaling, DNA repair, apoptotic processes and mitochondrial metabolism were found to be differentially expressed in endometrium. The findings of this study revealed proteins that hold strong candidature as potential drug targets to regulate the cellular processes implicating infertility and reduced receptivity of endometrium in women with polycystic ovary syndrome.
Assuntos
Proteínas Sanguíneas/análise , Endométrio/metabolismo , Infertilidade Feminina/metabolismo , Síndrome do Ovário Policístico/metabolismo , Proteínas/metabolismo , Adulto , Endométrio/patologia , Feminino , Humanos , ProteômicaRESUMO
The study involves isolation of arsenic resistant bacteria from soil samples. The characterization of bacteria isolates was based on 16S rRNA gene sequences. The phylogenetic consanguinity among isolates was studied employing rpoB and gltX gene sequence. RAPD-PCR technique was used to analyze genetic similarity between arsenic resistant isolates. In accordance with the results Bacillus subtilis and Bacillus pumilus strains may exhibit extensive horizontal gene transfer. Arsenic resistant potency in Bacillus sonorensis and high arsenite tolerance in Bacillus pumilus strains was identified. The RAPD-PCR primer OPO-02 amplified a 0.5kb DNA band specific to B. pumilus 3ZZZ strain and 0.75kb DNA band specific to B. subtilis 3PP. These unique DNA bands may have potential use as SCAR (Sequenced Characterized Amplified Region) molecular markers for identification of arsenic resistant B. pumilus and B. subtilis strains.
Assuntos
Arsênio/toxicidade , Bacillus pumilus/efeitos dos fármacos , Bacillus pumilus/genética , Bacillus subtilis/efeitos dos fármacos , Bacillus subtilis/genética , Reação em Cadeia da Polimerase/métodos , Técnica de Amplificação ao Acaso de DNA Polimórfico/métodos , Bacillus pumilus/isolamento & purificação , Bacillus subtilis/isolamento & purificação , Evolução Molecular , Genes Bacterianos , Concentração de Íons de Hidrogênio , Funções Verossimilhança , Filogenia , SoloRESUMO
The outbreak of coronavirus disease 2019 (COVID-19) has resulted in a world-wide crisis. To contain the virus, it is important to find infected individuals and isolate them to stop transmission. Various diagnostic techniques are used to check for infection. With the havoc that severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has created, it is imperative to work on alternative diagnostic techniques that can be used at both point of care with little or no expertise and at mass testing (i.e., when screening). Despite extensive research, to this date no specific effective treatment or cure is available to neutralize this viral infection. Globally, researchers are working to develop effective treatments, and several vaccines have been approved for public use. We found the studies that we explored for this review using appropriate key words for indexing in PubMed and Google Scholar from 2019 to 2020. We compile various techniques that have been used worldwide to diagnose and treat SARS-CoV-2 and discuss novel methods that may be modified for use in diagnosis and treatment. It is crucial to develop a more specific serological test for diagnosis that can rule out the possibility of COVID-19 and be used for mass testing. An affordable, safe, targeted, effective treatment must be developed to cure this disease, which has created a public health emergency of international concern.
Assuntos
Teste para COVID-19/tendências , COVID-19/diagnóstico , COVID-19/terapia , Saúde Global , Pneumonia Viral/diagnóstico , Pneumonia Viral/terapia , Vacinas contra COVID-19 , Humanos , Pandemias , SARS-CoV-2RESUMO
Early detection of breast cancer can be best facilitated by the development of precancerous markers. Serum proteins being the sensitive signatures, can be the ideal choice. We previously demonstrated the reduced levels of two serum proteins at a very early stage of tumorigenesis in a breast cancer model, developed in Wistar rats by 7,12-dimethylbenz[a]anthracene (DMBA) administration. Here we report the dysregulation of three more proteins in the serum collected at another early stage (15 weeks) of tumorigenesis in the same model. The proteins were identified (as Alpha-1-inhibitor III (Mug1), Immunoglobulin heavy chain variable region (IGHV), and Hypertrophied skeletal muscle protein GTF3) by MALDI-TOF MS after the screening and fingerprinting of serum samples by one-dimensional (1D) and two-dimensional (2D) electrophoresis respectively. Relative expression analysis of corresponding genes was also carried out, and the results were found as supporting the proteomic findings. In addition, the candidate proteins of the study and their corresponding ribonucleic acids (RNAs) were subjected to homology modelling and docking (using softwares like MODELLER, 3dRNA, Autodock4.0, and GROMACS etc), which revealed the binding sites for carcinogen (DMBA) and its nature of interaction with proteins and RNAs. Moreover, the network analysis by GeneMANIA unraveled the protein/gene functional network in which Mug1, IGHV, and GTF3 are involved. Based on the significant protein and gene expression alterations in early tumorigenesis, these proteins may prove very effective in search for biomarkers for the early detection of mammary cancer. Further, these proteins can also be tried as targets for chemotherapy.
Assuntos
Proteínas de Fase Aguda/metabolismo , Carcinogênese/metabolismo , Cadeias Pesadas de Imunoglobulinas/sangue , Região Variável de Imunoglobulina/sangue , Neoplasias Mamárias Experimentais/metabolismo , Transativadores/sangue , Animais , Biomarcadores Tumorais/sangue , Carcinoma/metabolismo , Detecção Precoce de Câncer , Feminino , Ratos , Ratos WistarRESUMO
Breast cancer is a major global health concern, appealing for precise prognostic approaches. Thus, the need is to have studies focusing on the identification and recognition of preliminary events leading to the disease. The present study reports the tracing of precancerous progression and serum proteomic analysis in a breast cancer model developed as a result of 7,12-dimethylbenz[a]anthracene (DMBA) administration. Mammary gland histological changes of prime importance were examined by histopathology, and immunohistochemical analysis with Ki-67 was performed to monitor enhanced cell proliferation, right from the onset of hyperplasia till neoplasia. Serum proteomics (one-dimensional (1D) and two-dimensional (2D) electrophoresis, followed by MALDI-TOF MS characterization) was performed to decipher the differentially expressed serum proteins in animals undergoing tumorigenesis vis-à-vis controls. The significance of our study lies in reporting the significantly reduced expression of two proteins: histone-lysine N-methyltransferase (SETD2) and sorting nexin-9 (SNX9) at very early stage (13 weeks) of tumorigenesis, while the full-fledged tumors developed after 6 months. The reduced expression of SETD2 and SNX9 was validated by western blotting and relative expression analysis using quantitative real-time PCR. These proteins may hence prove as potentially useful tools in search for prognostic markers for the early detection of mammary cancer.
Assuntos
Carcinogênese/patologia , Histona-Lisina N-Metiltransferase/biossíntese , Glândulas Mamárias Animais/patologia , Neoplasias Mamárias Animais/patologia , Nexinas de Classificação/biossíntese , 9,10-Dimetil-1,2-benzantraceno/toxicidade , Animais , Carcinogênese/induzido quimicamente , Proliferação de Células/efeitos dos fármacos , Feminino , Neoplasias Mamárias Animais/induzido quimicamente , Proteômica , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase em Tempo Real , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
OBJECTIVES: Cell free fetal DNA (cffDNA) and its hypermethylated RASSF1A gene signify a recent advancement in non-invasive prenatal diagnosis of feto-placental anomalies like pre-eclampsia. The study uses hypermethylated RASSF1A gene to quantify cffDNA and to assess its relationship with placental and urine proteins in pre-eclampsia cases. DESIGN AND METHODS: DNA was isolated from plasma samples of clinically diagnosed cases of pre-eclampsia (n=103) and normal pregnancy (n=616) from 21weeks of gestation. Through methylation sensitive enzyme (BstUI) digestion; followed by real time-polymerase chain reaction (RT-PCR), quantification of hypermethylated RASSF1A was done. Immunoassays determined: placental protein-13 (pp-13) and pregnancy associated plasma protein A (PAPP-A) and pyrogallol red molybdate assay for 24h urine protein. RESULTS: Highly significant differences between control and pre-eclampsia cases for hypermethylated RASSF1A concentrations were found; Group I: 33±7.35 vs 74.46±16.71, Group II: 53.75±16.65 vs 244.22±35.68, Group III: 93.25±19.08 vs 412.31±80.18, Group IV: 144.30±18.13 vs 1056.89±153.78, Group V: 307.55±40.76 vs 2763.76±259.76copies/ml. Multivariate Pearson's correlation analysis of hypermethylated RASSF1A with pp-13, PAPP-A and urine proteins showed positive and very highly significant (P<0.001) associations. CONCLUSIONS: Diagnostic potential of fetal specific, hypermethylated RASSF1A was evaluated. Its positive relationship with placental and urine proteins submit the case for considering it as a reliable marker for pre-eclampsia.
Assuntos
Metilação de DNA , Pré-Eclâmpsia/sangue , Pré-Eclâmpsia/diagnóstico , Diagnóstico Pré-Natal/métodos , Proteínas Supressoras de Tumor/genética , Adulto , Estudos de Casos e Controles , Feminino , Feto/metabolismo , Galectinas/sangue , Humanos , Gravidez , Proteínas da Gravidez/sangue , Proteína Plasmática A Associada à Gravidez/metabolismo , Proteinúria/urina , Adulto JovemRESUMO
Detection of human papillomavirus (HPV) types 6, 11, 16, 18, and 33 including co-infections among females attending gynaecological outpatient department and cancer clinics, was done by restriction fragment length polymorphism (using Rsa-1), of approximately 450bp amplicon, obtained by the amplification of the L1 region of HPV genome with consensus primers MY09/11 [Cancer Cells 7 (1989) 209]. The results were further tested with HPV type specific primers [J. Med. Virol. 29 (1989) 20]. The technique was found to be low-cost and less time consuming. The advantage of Rsa 1 over other enzymes was that it detects the five most prevalent HPV types commonly associated with warts, cervical dysplasia, and cancer.
Assuntos
Genoma Viral , Papillomaviridae/classificação , Papillomaviridae/genética , Infecções por Papillomavirus/diagnóstico , Polimorfismo de Fragmento de Restrição , Bacteriófago phi X 174/genética , Sequência de Bases , Primers do DNA , Desoxirribonucleases de Sítio Específico do Tipo II , Feminino , Globinas/genética , Humanos , Papillomaviridae/isolamento & purificação , Mapeamento por Restrição , Infecções Sexualmente Transmissíveis/diagnóstico , Infecções Sexualmente Transmissíveis/virologiaRESUMO
Methylenetetrahydrofolate reductase (MTHFR) gene located on chromosome 1p36.3 catalyses the conversion of 5,10-methylenetetrahydrofolate to 5,methyltetrahydrofolate, the major methyl donor for the conversion of homocysteine to methionine. Two common polymorphisms in the MTHFR gene have been identified, 677C>T in exon 4, leading to substitution of alanine by valine and 1298A>C in exon 7 which leads to the replacement of glutamic acid by alanine resulting into reduced enzyme activity. The potential influence of MTHFR activity on DNA methylation and on the availability of uridylates and thymidylates for DNA synthesis and repair makes MTHFR an attractive candidate for cancer predisposing gene. In order to elucidate the role of MTHFR polymorphism in cervical cancer, both the exons for 677C>T and 1298A>C mutations were analyzed among 219 females, including 77 females with normal cervical cytology, 80 with cervical dysplasia and 62 with squamous cell carcinoma of uterine cervix. Females with mutant allele at 677 position (CT/TT genotypes) were found to be almost three times the risk of cervical dysplasia than females with CC genotype [OR, 2.9; (CI, 1.5-5.7)], but were less likely to develop squamous cell carcinoma [OR, 1.5 (CI, 0.7-3.2)]. Similar findings were observed for mutation at 1298 position, females with AC/CC genotypes were almost four times the risk of cervical dysplasia [OR, 4.3 (CI, 2.1-9.0)], as compared to AA genotype. Our study lends further support to the hypothesis that the MTHFR polymorphism (677C>T or 1298A>C) is involved in susceptibility to cervical dysplasia.