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1.
J Med Life ; 13(2): 241-248, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32742521

RESUMO

Cell culture is one of the most commonly used techniques in the production of biological products. Many physical and chemical parameters may affect cell growth and proliferation. This study was conducted to investigate the effect of chemical components as supplements using the experimental design method, which aimed at reducing the number of experiments. For this purpose, supplements including chemical components using four levels, with three replications in suspension and batch culture conditions, were examined for 72 hours using the Taguchi experimental design method. From the experiments, it was concluded that the culture media composition had a significant impact on final cell count and pH. High concentrations of different media composition alone were insufficient to ensure higher cell count. According to the results, this insufficiency was associated with an increase of 20% in the number of final cells. In the majority of cultures, the number of final cells at 48 hours increased relative to the number of final cells at 24 hours after culturing the cells.


Assuntos
Técnicas de Cultura de Células/métodos , Vírus da Febre Aftosa/imunologia , Rim/citologia , Vacinas Virais/imunologia , Aminoácidos/farmacologia , Animais , Contagem de Células , Células Cultivadas , Cricetinae , Vírus da Febre Aftosa/efeitos dos fármacos , Glucose/farmacologia , Concentração de Íons de Hidrogênio , Polietilenoglicóis/química , Proteínas/farmacologia , Vitaminas/farmacologia
2.
Zhongguo Shi Yan Xue Ye Xue Za Zhi ; 28(4): 1357-1362, 2020 Aug.
Artigo em Chinês | MEDLINE | ID: mdl-32798426

RESUMO

OBJECTIVE: To explore the method for inducing the differentiation of bone marrow cells into megakaryocytes in vitro so as to use for evaluating the activity of traditional Chinese medicines. METHODS: The bone marrow cells were separated from femurs and tibias of mice. The experiments were divided into 4 groups: control (no adding cytokines), TPO (adding 50 ng/ml TPO), TPO+SCF (50 ng/ml+50 ng/ml) and TPO+SCF+IL-6+IL-9 (50 ng/ml+50 ng/ml+20 ng/ml+20 ng/ml). The bone marrow cells in 4 groups were cultured in vitro for 6 d. Then the cell growth status was observed by the inverted microscopy, and the cell count was detected by using the automatic cell counter. The ratio and absolute count of megakaryocytes were detected by flow cytometry. RESULTS: Compared with control, three induction methods could stimulate the differentiation of bone marrow cells into megakaryocytes in vitro. TPO could slightly enhance the differentiation of bone marrow cells into megakaryocytes. Both the combination of TPO and SCF, and the combination of TPO, SCF, IL-6 and IL-9 could intensively stimulate proliferation of bone morrow cells and promote the differentiation of bone marrow cells into megakaryocytes. The addition of IL-6 and IL-9 could decrease the proliferation of non-megakaryocytes, but promote the differentiation of bone marrow cells into megakaryocytes. CONCLUSION: The optimized differentiation of bone marrow cells into megakaryocytes has been completed by co-induction regimen of TPO, SCF, IL-6 and IL-9, which can be used to screen and evaluate traditional Chinese medicines promoting formation of platelets.


Assuntos
Interleucina-3 , Megacariócitos , Animais , Células da Medula Óssea , Contagem de Células , Diferenciação Celular , Divisão Celular , Células Cultivadas , Camundongos , Fator de Células-Tronco , Trombopoetina
3.
Int J Nanomedicine ; 15: 4441-4452, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32606688

RESUMO

Purpose: The present study focuses on threshold levels for cytotoxicity after long-term and repetitive exposure for HUVEC as a model for the specific microvascular endothelial system. Furthermore, possible genotoxic effects and functional impairment caused by ZnO NPs in HUVEC are elucidated. Methods: Thresholds for cytotoxic effects are determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and Annexin V assay. To demonstrate DNA damage, single-cell microgel electrophoresis (comet) assay is performed after exposure to sub-cytotoxic concentrations of ZnO NPs. The proliferation assay, dot blot assay and capillary tube formation assay are also carried out to analyze functional impairment. Results: NPs showed to be spherical in shape with an average size of 45-55 nm. Long-term exposure as well as repetitive exposure with ZnO NPs exceeding 25 µg/mL lead to decreased viability in HUVEC. In addition, DNA damage was indicated by the comet assay after long-term and repetitive exposure. Twenty-four hours after long-term exposure, the proliferation assay does not show any difference between negative control and exposed cells. Forty-eight hours after exposure, HUVEC show an inverse concentration-related ability to proliferate. The dot blot assay provides evidence that ZnO NPs lead to a decreased release of VEGF, while capillary tube formation assay shows restriction in the ability of HUVEC to build tubes and meshes as a first step in angiogenesis. Conclusion: Sub-cytotoxic concentrations of ZnO NPs lead to DNA damage and functional impairment in HUVEC. Based on these data, ZnO NPs may affect neo-angiogenesis. Further investigation based on tissue cultures is required to elucidate the impact of ZnO NPs on human cell systems.


Assuntos
Dano ao DNA , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Nanopartículas Metálicas/toxicidade , Óxido de Zinco/toxicidade , Anexina A5/metabolismo , Apoptose/efeitos dos fármacos , Contagem de Células , Sobrevivência Celular/efeitos dos fármacos , Humanos , Neovascularização Fisiológica/efeitos dos fármacos
4.
PLoS One ; 15(7): e0223395, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32645024

RESUMO

Development of the cerebral cortex may be influenced by the composition of the maternal gut microbiota. To test this possibility, we administered probiotic Lactococcus lactis in drinking water to mouse dams from day 10.5 of gestation until pups reached postnatal day 1 (P1). Pups were assessed in a battery of behavioral tests starting at 10 weeks old. We found that females, but not males, exposed to probiotic during prenatal development spent more time in the center of the open field and displayed decreased freezing time in cue associated learning, compared to controls. Furthermore, we found that probiotic exposure changed the density of cortical neurons and increased the density of blood vessels in the cortical plate of P1 pups. Sex-specific differences were observed in the number of mitotic neural progenitor cells, which were increased in probiotic exposed female pups. In addition, we found that probiotic treatment in the latter half of pregnancy significantly increased plasma oxytocin levels in mouse dams, but not in the offspring. These results suggest that exposure of naïve, unstressed dams to probiotic may exert sex-specific long-term effects on cortical development and anxiety related behavior in the offspring.


Assuntos
Ansiedade/prevenção & controle , Córtex Cerebral/efeitos dos fármacos , Lactococcus lactis , Efeitos Tardios da Exposição Pré-Natal/psicologia , Probióticos/farmacologia , Animais , Animais Recém-Nascidos , Contagem de Células , Córtex Cerebral/irrigação sanguínea , Córtex Cerebral/citologia , Córtex Cerebral/crescimento & desenvolvimento , Medo , Feminino , Aprendizagem , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/citologia , Ocitocina/metabolismo , Gravidez , Caracteres Sexuais
5.
PLoS One ; 15(7): e0232559, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32658922

RESUMO

PRESENILIN 2 (PSEN2) is one of the genes mutated in early onset familial Alzheimer's disease (EOfAD). PSEN2 shares significant amino acid sequence identity with another EOfAD-related gene PRESENILIN 1 (PSEN1), and partial functional redundancy is seen between these two genes. However, the complete range of functions of PSEN1 and PSEN2 is not yet understood. In this study, we performed targeted mutagenesis of the zebrafish psen2 gene to generate a premature termination codon close downstream of the translation start with the intention of creating a null mutation. Homozygotes for this mutation, psen2S4Ter, are viable and fertile, and adults do not show any gross psen2-dependent pigmentation defects, arguing against significant loss of γ-secretase activity. Also, assessment of the numbers of Dorsal Longitudinal Ascending (DoLA) interneurons that are responsive to psen2 but not psen1 activity during embryogenesis did not reveal decreased psen2 function. Transcripts containing the S4Ter mutation show no evidence of destabilization by nonsense-mediated decay. Forced expression in zebrafish embryos of fusions of psen2S4Ter 5' mRNA sequences with sequence encoding enhanced green fluorescent protein (EGFP) indicated that the psen2S4Ter mutation permits utilization of cryptic, novel downstream translation start codons. These likely initiate translation of N-terminally truncated Psen2 proteins lacking late endosomal/lysosomal localization sequences and that obey the "reading frame preservation rule" of PRESENILIN EOfAD mutations. Transcriptome analysis of entire brains from a 6-month-old family of wild type, heterozygous and homozygous psen2S4Ter female siblings revealed profoundly dominant effects on gene expression likely indicating changes in ribosomal, mitochondrial, and anion transport functions.


Assuntos
Códon de Terminação/genética , Perfilação da Expressão Gênica , Mitocôndrias/genética , Mutação , Presenilina-2/genética , Ribossomos/genética , Proteínas de Peixe-Zebra/genética , Alelos , Animais , Contagem de Células , Homozigoto , Hipóxia/genética , Neurônios/citologia , Estabilidade de RNA/genética , Peixe-Zebra/embriologia , Peixe-Zebra/genética
6.
Anticancer Res ; 40(8): 4701-4706, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32727795

RESUMO

BACKGROUND/AIM: Bovine mastitis is caused by the invasion and propagation of pathogenic microorganisms into the udder and mammary gland tissues of cattle. In this study, the therapeutic effect of a low-molecular-weight whey protein (LMW-WP) on bovine mastitis was evaluated. MATERIALS AND METHODS: LMW-WP was orally, intraperitoneally, and vaginally administered to bovine with mastitis. The number of somatic cells in milk was measured 24 h before the administration of LMW-WP. The effect of LMW-WP on cytokine production was measured with a microarray that evaluates the expression of cytokines. RESULTS: In the group that received 1,000 mg intraperitoneally, the somatic cell count was reduced to less than 400,000 at the shipment standard value in three of the four udders, indicating 75% efficacy. The group that received 1,000 mg by vaginal administration showed 67% efficacy. It was confirmed that LMW-WP increased the production of cytokines such as IL-5, IL-6, IL-9, IL-12, MCP-1, and VEGF in mouse macrophage cells, but it did not show any antibacterial activity. CONCLUSION: LMW-WP may be an effective therapeutic agent for bovine mastitis.


Assuntos
Macrófagos/efeitos dos fármacos , Mastite Bovina/tratamento farmacológico , Proteínas do Soro do Leite/farmacologia , Animais , Antibacterianos/farmacologia , Bovinos , Contagem de Células/métodos , Linhagem Celular , Citocinas/metabolismo , Feminino , Glândulas Mamárias Animais/metabolismo , Mastite Bovina/microbiologia , Camundongos , Leite/metabolismo , Células RAW 264.7
7.
Ann Hematol ; 99(7): 1515-1523, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32506245

RESUMO

Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening systemic hyperinflammatory disorder. We found recently that repeated lipopolysaccharide (LPS) treatment induces HLH-like features in senescence-accelerated mice (SAMP1/TA-1) but not in senescence-resistant control mice (SAMR1). In this study, we analyzed the dynamics of hematopoiesis in this mouse model of HLH. When treated repeatedly with LPS, the numbers of myeloid progenitor cells (CFU-GM) and B-lymphoid progenitor cells (CFU-preB) in the bone marrow (BM) rapidly decreased after each treatment in both strains. The number of CFU-GM in SAMP1/TA-1 and SAMR1, and of CFU-preB in SAMR1, returned to pretreatment levels by 7 days after each treatment. However, the recovery in the number of CFU-preB in SAMP1/TA-1 was limited. In both strains, the BM expression of genes encoding positive regulators of myelopoiesis (granulocyte colony-stimulating factor (G-CSF), granulocyte macrophage colony-stimulating factor (GM-CSF), and interleukin (IL)-6), and negative regulators of B lymphopoiesis (tumor necrosis factor (TNF)-α) was increased. The expression of genes encoding positive regulators of B lymphopoiesis (stromal-cell derived factor (SDF)-1, IL-7, and stem cell factor (SCF)) was persistently decreased in SAMP1/TA-1 but not in SAMR1. Expression of the gene encoding p16INK4a and the proportion of ß-galactosidase-positive cells were increased in cultured stromal cells obtained from LPS-treated SAMP1/TA-1 but not in those from LPS-treated SAMR1. LPS treatment induced qualitative changes in stromal cells, which comprise the microenvironment supporting appropriate hematopoiesis, in SAMP1/TA-1; these stromal cell changes are inferred to disrupt the dynamics of hematopoiesis. Thus, hematopoietic tissue is one of the organs that suffer life-threatening damage in HLH.


Assuntos
Medula Óssea/patologia , Hematopoese/fisiologia , Células-Tronco Hematopoéticas/patologia , Linfo-Histiocitose Hemofagocítica/patologia , Linfo-Histiocitose Hemofagocítica/fisiopatologia , Nicho de Células-Tronco/fisiologia , Animais , Células da Medula Óssea/patologia , Células da Medula Óssea/fisiologia , Contagem de Células , Células Cultivadas , Microambiente Celular/fisiologia , Modelos Animais de Doenças , Células-Tronco Hematopoéticas/fisiologia , Lipopolissacarídeos , Linfo-Histiocitose Hemofagocítica/induzido quimicamente , Masculino , Camundongos , Células Estromais/patologia
8.
Sheng Wu Gong Cheng Xue Bao ; 36(6): 1209-1215, 2020 Jun 25.
Artigo em Chinês | MEDLINE | ID: mdl-32597070

RESUMO

Bioreactors have been central in monoclonal antibodies and vaccines manufacturing by mammalian cells in suspension culture. Numerical simulation of five impeller combinations in a stirred bioreactor was conducted, and characteristics of velocity vectors, distributions of gas hold-up, distributions of shear rate in the bioreactor using 5 impeller combinations were numerically elucidated. In addition, genetically engineered CHO cells were cultivated in bioreactor installed with 5 different impeller combinations in fed-batch culture mode. The cell growth and antibody level were directly related to the maximum shear rate in the bioreactor, and the highest viable cell density and the peak antibody level were achieved in FBMI3 impeller combination, indicating that CHO cells are sensitive to shear force produced by impeller movement when cells were cultivated in bioreactor at large scale, and the maximum shear rate would play key roles in scaling-up of bioreactor at industrial scale.


Assuntos
Técnicas de Cultura Celular por Lotes , Reatores Biológicos , Simulação por Computador , Microbiologia Industrial , Animais , Reatores Biológicos/normas , Células CHO , Contagem de Células , Cricetinae , Cricetulus , Microbiologia Industrial/instrumentação , Microbiologia Industrial/métodos
9.
Eur Urol Focus ; 6(5): 1124-1129, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32563676

RESUMO

BACKGROUND: Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), involves multiple organs. Testicular involvement is largely unknown. OBJECTIVE: To determine the pathological changes and whether SARS-CoV-2 can be detected in the testes of deceased COVID-19 patients. DESIGN, SETTING, AND PARTICIPANTS: Postmortem examination of the testes from 12 COVID-19 patients was performed using light and electron microscopy, and immunohistochemistry for lymphocytic and histiocytic markers. Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the virus in testicular tissue. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Seminiferous tubular injury was assessed as none, mild, moderate, or severe according to the extent of tubular damage. Leydig cells in the interstitium were counted in ten 400× microscopy fields. RESULTS AND LIMITATIONS: Microscopically, Sertoli cells showed swelling, vacuolation and cytoplasmic rarefaction, detachment from tubular basement membranes, and loss and sloughing into lumens of the intratubular cell mass. Two, five, and four of 11 cases showed mild, moderate, and severe injury, respectively. The mean number of Leydig cells in COVID-19 testes was significantly lower than in the control group (2.2 vs 7.8, p < 0.001). In the interstitium there was edema and mild inflammatory infiltrates composed of T lymphocytes and histiocytes. Transmission EM did not identify viral particles in three cases. RT-PCR detected the virus in one of 12 cases. CONCLUSIONS: Testes from COVID-19 patients exhibited significant seminiferous tubular injury, reduced Leydig cells, and mild lymphocytic inflammation. We found no evidence of SARS-CoV-2 virus in the testes in the majority (90%) of the cases by RT-PCR, and in none by electron microscopy. These findings can provide evidence-based guidance for sperm donation and inform management strategies to mitigate the risk of testicular injury during the COVID-19 disease course. PATIENT SUMMARY: We examined the testes of deceased COVID-19 patients. We found significant damage to the testicular parenchyma. However, virus was not detected in testes in the majority of cases.


Assuntos
Infecções por Coronavirus/patologia , Pneumonia Viral/patologia , Túbulos Seminíferos/patologia , Testículo/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Betacoronavirus , Contagem de Células , Infecções por Coronavirus/metabolismo , Infecções por Coronavirus/fisiopatologia , Humanos , Inflamação , Células Intersticiais do Testículo/patologia , Células Intersticiais do Testículo/ultraestrutura , Masculino , Microscopia Eletrônica , Pessoa de Meia-Idade , Pandemias , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/metabolismo , Pneumonia Viral/fisiopatologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Túbulos Seminíferos/ultraestrutura , Células de Sertoli/patologia , Células de Sertoli/ultraestrutura , Espermatogênese/fisiologia , Testículo/metabolismo , Testículo/ultraestrutura , Testículo/virologia
10.
PLoS One ; 15(6): e0234202, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32525919

RESUMO

PURPOSE: To investigate the factors associated with endothelial survival after Descemet's membrane endothelial keratoplasty (DMEK) in eyes of Asian patients with bullous keratopathy (BK). METHODS: In this retrospective, consecutive interventional case series, 72 eyes of 72 patients who underwent DMEK were evaluated. Best corrected visual acuity (BCVA) and corneal endothelial cell density (ECD) were assessed at 12 months postoperatively. Multiple regression analysis was performed to assess parameters such as age, sex, axial length, preoperative visual acuity, re-bubbling, the ratio of graft to cornea area, iris damage scores, types of filling gases, air or SF6 volume in the anterior chamber (AC) on postoperative day 1, and ECD loss rates at 12 months postoperatively. RESULTS: BCVA improved significantly at 12 months after DMEK (P < .001). The rate of ECD loss at 12 months after DMEK was 54.4 ± 16.1%. Multiple linear regression analysis showed that a larger ratio of graft to corneal area (P = 0.0061) and higher donor ECD (P = 0.042) were the primary factors for a lower ECD loss rate at 12 months after DMEK. CONCLUSION: A relatively larger graft size compared to the host cornea and more donor ECD might help endothelial survival in patients with BK. Moreover, for such patients, the surgeon should attempt to use a relatively larger graft size when performing DMEK, particularly in Asian eyes.


Assuntos
Doenças da Córnea/patologia , Lâmina Limitante Posterior/patologia , Células Endoteliais/patologia , Adulto , Idoso , Ásia , Contagem de Células , Sobrevivência Celular , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos
11.
Zhejiang Da Xue Xue Bao Yi Xue Ban ; 49(2): 220-226, 2020 May 25.
Artigo em Chinês | MEDLINE | ID: covidwho-233220

RESUMO

OBJECTIVE: To investigate the effect of corticosteroids therapy on the inflammatory response in a critically ill coronavirus disease 2019 (COVID-19) patient. METHODS: A 55-year old female patient with critical ill COVID-19 was admitted in Taizhou Hospital on January 19, 2020. The patient was treated with methylprednisolone 80 mg on the 2nd day after admission. Thereafter, the dose was adjusted in a timely manner and the therapy lasted for 13 days. The peripheral lymphocyte subsets (CD3+T, CD4+ T, CD8+ T, NK cells, B cells), as well as serum levels of lymphocyte factors (IL-2, IL-4, IL-6, IL-10, TNF-α, IFN-γ) were dynamically monitored. RESULTS: On D1 of admission, the numbers of peripheral blood CD3+ T, CD4+ T, CD8+ T, and NK cells were significantly lower than the normal range. With the improvement of the disease, the numbers of CD3+ T, CD8+ T and CD4 + T cells gradually recovered and showed a linear growth trend (linear fitting equation: Y=18.59X+109.4, P<0.05). On D2 of admission, the patient's IL-6 and IL-10 levels were significantly higher than normal values, IFN-γ was at a normal high value, and then rapidly decreased; IL-2, IL-4, and TNF-α were all in the normal range. On the D6 and D7, the IL-6 and IL-10 decreased to the normal range for the first time. On the D18, the sputum virus nucleic acid test was negative for the first time, and the fecal virus nucleic acid test was still positive; on the D20 the sputum and fecal virus nucleic acid test were both negative. On D34, the patient recovered and was discharged. At the discharge the muscle strength score of the patient was 44 and the daily life ability evaluation was 90. CONCLUSIONS: In the absence of effective antiviral drugs, early use of appropriate doses of corticosteroids in critically ill patient with COVID-19 can quickly alleviate inflammatory response and improve clinical symptoms, however, it may reduce the number of T cells, and to adjust the dose in time is necessary.


Assuntos
Betacoronavirus , Infecções por Coronavirus , Metilprednisolona , Pandemias , Pneumonia Viral , Betacoronavirus/isolamento & purificação , Contagem de Células , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/fisiopatologia , Estado Terminal , Citocinas/sangue , Feminino , Humanos , Metilprednisolona/administração & dosagem , Metilprednisolona/efeitos adversos , Pessoa de Meia-Idade , Pneumonia Viral/diagnóstico , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/imunologia , Pneumonia Viral/fisiopatologia , Subpopulações de Linfócitos T/efeitos dos fármacos , Resultado do Tratamento
12.
Clin Chim Acta ; 508: 98-102, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32405079

RESUMO

BACKGROUND: Novel coronavirus infectious disease (COVID-19) has been spreading worldwide, and tracking laboratory indexes during the diagnosis and treatment of patients with severe COVID-19 can provide a reference for patients in other countries and regions. METHODS: We closely tracked the epidemiological history, diagnosis and treatment process, as well as dynamic changes in routine blood indicators, of a severe COVID-19 patient who was hospitalized for 26 days. RESULTS: Our study found that the patient's condition worsened in the first week after admission, white blood cells (WBCs), neutrophils, lymphocytes, monocytes, eosinophils, red blood cells (RBCs), hemoglobin, neutrophil lymphocyte ratio (NLR), platelets (PLT) and platelet lymphocyte ratio (PLR) decreased. On the 7th day of admission, the levels of these cells decreased to their lowest values, though the red blood cell distribution width (RDW) and C-reactive protein (CRP) level remained at high values. From 8 to 14 days of admission, the patient's condition improved, hypoxemia was corrected, and mechanical ventilation was discontinued. The number of WBCs, neutrophils, monocytes, eosinophils and lymphocytes increased gradually, and the erythrocyte parameters stopped declining and stabilized in a certain range; CRP decreased rapidly. On the 20th day of admission, the nucleic acid test was negative, WBC, neutrophil, CRP, NLR and PLR decreased gradually, and monocyte, lymphocyte, and eosinophil counts increased. Although RBCs and hemoglobin (Hb) levels continued to decrease, RDW gradually increased, indicating the recovery of hematopoiesis. In addition, it should be noted that monocytes and eosinophils were at extremely low levels within 10 days after admission; the recovery time of eosinophils was approximately 12 days after admission, which was earlier than other parameters, which might be of great value in judging the progress of the disease. CONCLUSIONS: Dynamic changes in routine blood parameters might be helpful for the prognosis of COVID-19 patients and evaluation of the treatment effect.


Assuntos
Betacoronavirus/patogenicidade , Infecções por Coronavirus/sangue , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/sangue , Pneumonia Viral/diagnóstico , Antibacterianos/uso terapêutico , Betacoronavirus/efeitos dos fármacos , Biomarcadores/sangue , Plaquetas/efeitos dos fármacos , Plaquetas/patologia , Plaquetas/virologia , Proteína C-Reativa/metabolismo , Contagem de Células , Convalescença , Infecções por Coronavirus/fisiopatologia , Infecções por Coronavirus/terapia , Eritrócitos/efeitos dos fármacos , Eritrócitos/patologia , Eritrócitos/virologia , Feminino , Humanos , Pessoa de Meia-Idade , Monócitos/efeitos dos fármacos , Monócitos/patologia , Monócitos/virologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/patologia , Neutrófilos/virologia , Oseltamivir/uso terapêutico , Pandemias , Pneumonia Viral/fisiopatologia , Pneumonia Viral/terapia , Prognóstico , Respiração Artificial , Índice de Gravidade de Doença
13.
Clin Chim Acta ; 508: 122-129, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32417210

RESUMO

BACKGROUND: The underlying changes of peripheral blood inflammatory cells (PBICs) in COVID-19 patients are little known. Moreover, the risk factors for the underlying changes of PBICs and their predicting role in severe COVID-19 patients remain uncertain. MATERIAL AND METHODS: This retrospective study including two cohorts: the main cohort enrolling 45 patients of severe type serving as study group, and the secondary cohort enrolling 12 patients of no-severe type serving as control group. The PBICs analysis was based on blood routine and lymphocyte subsets. The inflammatory cell levels were compared among patients according to clinical classifications, disease-associated phases, as well as one-month outcomes. RESULTS: Compared with patients of non-severe type, the patients of severe type suffered from significantly decreased counts of lymphocytes, eosinophils, basophils, but increased counts of neutrophils. These PBICs alterations got improved in recovery phase, but persisted or got worse in aggravated phase. Compared with patients in discharged group, the patients in un-discharged/died group suffered from decreased counts of total T lymphocytes, CD4 + T lymphocytes, CD8 + T lymphocytes, as well as NK cells at 2 weeks after treatment. Clinical classification-critically severe was the independently risk factor for lymphopenia (OR = 7.701, 95%CI:1.265-46.893, P = 0.027), eosinopenia (OR = 5.595, 95%CI:1.008-31.054, P = 0.049), and worse one-month outcome (OR = 8.984; 95%CI:1.021-79.061, P = 0.048). CONCLUSION: Lymphopenia and eosinopenia may serve as predictors of disease severity and disease progression in COVID-19 patients, and enhancing the cellular immunity may contribute to COVID-19 treatment. Thus, PBICs might become a sentinel of COVID-19, and it deserves attention during COVID-19 treatment.


Assuntos
Betacoronavirus/patogenicidade , Infecções por Coronavirus/diagnóstico , Eosinófilos/patologia , Subpopulações de Linfócitos/patologia , Linfopenia/diagnóstico , Pneumonia Viral/diagnóstico , Idoso , Biomarcadores/sangue , Contagem de Células , Infecções por Coronavirus/sangue , Infecções por Coronavirus/fisiopatologia , Infecções por Coronavirus/virologia , Progressão da Doença , Eosinófilos/virologia , Feminino , Humanos , Células Matadoras Naturais/patologia , Células Matadoras Naturais/virologia , Subpopulações de Linfócitos/virologia , Linfopenia/sangue , Linfopenia/fisiopatologia , Linfopenia/virologia , Masculino , Pessoa de Meia-Idade , Monócitos/patologia , Monócitos/virologia , Neutrófilos/patologia , Neutrófilos/virologia , Pandemias , Pneumonia Viral/sangue , Pneumonia Viral/fisiopatologia , Pneumonia Viral/virologia , Prognóstico , Estudos Retrospectivos , Índice de Gravidade de Doença , Análise de Sobrevida
14.
Zhejiang Da Xue Xue Bao Yi Xue Ban ; 49(2): 220-226, 2020 May 25.
Artigo em Chinês | MEDLINE | ID: mdl-32391668

RESUMO

OBJECTIVE: To investigate the effect of corticosteroids therapy on the inflammatory response in a critically ill coronavirus disease 2019 (COVID-19) patient. METHODS: A 55-year old female patient with critical ill COVID-19 was admitted in Taizhou Hospital on January 19, 2020. The patient was treated with methylprednisolone 80 mg on the 2nd day after admission. Thereafter, the dose was adjusted in a timely manner and the therapy lasted for 13 days. The peripheral lymphocyte subsets (CD3+T, CD4+ T, CD8+ T, NK cells, B cells), as well as serum levels of lymphocyte factors (IL-2, IL-4, IL-6, IL-10, TNF-α, IFN-γ) were dynamically monitored. RESULTS: On D1 of admission, the numbers of peripheral blood CD3+ T, CD4+ T, CD8+ T, and NK cells were significantly lower than the normal range. With the improvement of the disease, the numbers of CD3+ T, CD8+ T and CD4 + T cells gradually recovered and showed a linear growth trend (linear fitting equation: Y=18.59X+109.4, P<0.05). On D2 of admission, the patient's IL-6 and IL-10 levels were significantly higher than normal values, IFN-γ was at a normal high value, and then rapidly decreased; IL-2, IL-4, and TNF-α were all in the normal range. On the D6 and D7, the IL-6 and IL-10 decreased to the normal range for the first time. On the D18, the sputum virus nucleic acid test was negative for the first time, and the fecal virus nucleic acid test was still positive; on the D20 the sputum and fecal virus nucleic acid test were both negative. On D34, the patient recovered and was discharged. At the discharge the muscle strength score of the patient was 44 and the daily life ability evaluation was 90. CONCLUSIONS: In the absence of effective antiviral drugs, early use of appropriate doses of corticosteroids in critically ill patient with COVID-19 can quickly alleviate inflammatory response and improve clinical symptoms, however, it may reduce the number of T cells, and to adjust the dose in time is necessary.


Assuntos
Betacoronavirus , Infecções por Coronavirus , Metilprednisolona , Pandemias , Pneumonia Viral , Betacoronavirus/isolamento & purificação , Contagem de Células , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/fisiopatologia , Estado Terminal , Citocinas/sangue , Feminino , Humanos , Metilprednisolona/administração & dosagem , Metilprednisolona/efeitos adversos , Pessoa de Meia-Idade , Pneumonia Viral/diagnóstico , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/imunologia , Pneumonia Viral/fisiopatologia , Subpopulações de Linfócitos T/efeitos dos fármacos , Resultado do Tratamento
15.
PLoS Comput Biol ; 16(5): e1007611, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32379821

RESUMO

Modeling cancer cells is essential to better understand the dynamic nature of brain tumors and glioma cells, including their invasion of normal brain. Our goal is to study how the morphology of the glioma cell influences the formation of patterns of collective behavior such as flocks (cells moving in the same direction) or streams (cells moving in opposite direction) referred to as oncostream. We have observed experimentally that the presence of oncostreams correlates with tumor progression. We propose an original agent-based model that considers each cell as an ellipsoid. We show that stretching cells from round to ellipsoid increases stream formation. A systematic numerical investigation of the model was implemented in [Formula: see text]. We deduce a phase diagram identifying key regimes for the dynamics (e.g. formation of flocks, streams, scattering). Moreover, we study the effect of cellular density and show that, in contrast to classical models of flocking, increasing cellular density reduces the formation of flocks. We observe similar patterns in [Formula: see text] with the noticeable difference that stream formation is more ubiquitous compared to flock formation.


Assuntos
Neoplasias Encefálicas/patologia , Biologia Computacional/métodos , Glioma/patologia , Contagem de Células/métodos , Movimento Celular/fisiologia , Forma Celular/fisiologia , Humanos , Modelos Biológicos , Modelos Teóricos , Simulação de Dinâmica Molecular
16.
BMC Bioinformatics ; 21(1): 187, 2020 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-32408861

RESUMO

BACKGROUND: Cardiotoxicity, characterized by severe cardiac dysfunction, is a major problem in patients treated with different classes of anticancer drugs. Development of predictable human-based models and assays for drug screening are crucial for preventing potential drug-induced adverse effects. Current animal in vivo models and cell lines are not always adequate to represent human biology. Alternatively, human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) show great potential for disease modelling and drug-induced toxicity screenings. Fully automated high-throughput screening of drug toxicity on hiPSC-CMs by fluorescence image analysis is, however, very challenging, due to clustered cell growth patterns and strong intracellular and intercellular variation in the expression of fluorescent markers. RESULTS: In this paper, we report on the development of a fully automated image analysis system for quantification of cardiotoxic phenotypes from hiPSC-CMs that are treated with various concentrations of anticancer drugs doxorubicin or crizotinib. This high-throughput system relies on single-cell segmentation by nuclear signal extraction, fuzzy C-mean clustering of cardiac α-actinin signal, and finally nuclear signal propagation. When compared to manual segmentation, it generates precision and recall scores of 0.81 and 0.93, respectively. CONCLUSIONS: Our results show that our fully automated image analysis system can reliably segment cardiomyocytes even with heterogeneous α-actinin signals.


Assuntos
Cardiotoxicidade/patologia , Processamento de Imagem Assistida por Computador , Células-Tronco Pluripotentes Induzidas/patologia , Miócitos Cardíacos/patologia , Automação , Comunicação Celular , Contagem de Células , Linhagem Celular , Doxorrubicina/efeitos adversos , Humanos , Fenótipo
17.
PLoS Genet ; 16(5): e1008782, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32421721

RESUMO

The planar cell polarity pathway is required for heart development and whilst the functions of most pathway members are known, the roles of the jnk genes in cardiac morphogenesis remain unknown as mouse mutants exhibit functional redundancy, with early embryonic lethality of compound mutants. In this study zebrafish were used to overcome early embryonic lethality in mouse models and establish the requirement for Jnk in heart development. Whole mount in-situ hybridisation and RT-PCR demonstrated that evolutionarily conserved alternative spliced jnk1a and jnk1b transcripts were expressed in the early developing heart. Maternal zygotic null mutant zebrafish lines for jnk1a and jnk1b, generated using CRISPR-Cas9, revealed a requirement for jnk1a in formation of the proximal, first heart field (FHF)-derived portion of the cardiac ventricular chamber. Rescue of the jnk1a mutant cardiac phenotype was only possible by injection of the jnk1a EX7 Lg alternatively spliced transcript. Analysis of mutants indicated that there was a reduction in the size of the hand2 expression field in jnk1a mutants which led to a specific reduction in FHF ventricular cardiomyocytes within the anterior lateral plate mesoderm. Moreover, the jnk1a mutant ventricular defect could be rescued by injection of hand2 mRNA. This study reveals a novel and critical requirement for Jnk1 in heart development and highlights the importance of alternative splicing in vertebrate cardiac morphogenesis. Genetic pathways functioning through jnk1 may be important in human heart malformations with left ventricular hypoplasia.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Ventrículos do Coração/citologia , Proteína Quinase 8 Ativada por Mitógeno/genética , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/embriologia , Processamento Alternativo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Contagem de Células , Células Cultivadas , Éxons , Regulação da Expressão Gênica no Desenvolvimento , Ventrículos do Coração/embriologia , Ventrículos do Coração/metabolismo , Proteína Quinase 8 Ativada por Mitógeno/metabolismo , Miócitos Cardíacos/citologia , Miócitos Cardíacos/metabolismo , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética
18.
Adv Exp Med Biol ; 1220: 1-10, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32304076

RESUMO

Breast cancer metastasis is a complex multistep process during which tumor cells undergo structural and functional changes that allow them to move away from the primary tumor and disseminate to distant organs and tissues. Despite the inefficiency of this process, some populations of circulating tumor cells (CTCs), which are those cells responsible of metastases formation, are able to survive in blood circulation and grow into secondary tumors. Metastatic breast cancer remains an incurable disease, and the phenomenon of metastasis represents the larger cause of death in these patients. The application of liquid biopsy techniques and the advancements in the field have shown the prognostic value of CTCs, suggesting the importance that CTCs analyses may have in the clinic. However, their implementation in routine clinic has not been yet achieved due to the yet small body of evidence showing their clinical utility. This introductory chapter will revise the key aspects of breast cancer metastasis and discuss the importance of CTC analyses in the management of breast cancer patients.


Assuntos
Neoplasias da Mama/patologia , Metástase Neoplásica , Células Neoplásicas Circulantes/patologia , Contagem de Células , Humanos
19.
Adv Exp Med Biol ; 1220: 45-59, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32304079

RESUMO

The majority of deaths related to breast cancer are caused by metastasis. Understanding the process of metastasis is key to achieve a reduction on breast cancer mortality. Currently, liquid biopsies are gaining attention in this regard. Circulating tumor cells (CTCs), an important component of liquid biopsies, are cells shed from primary tumor that disseminate to blood circulation being responsible of distal metastasis. Hence, the study CTCs is a promising alternative to monitor the progress of metastasis disease and can be used for early diagnosis of cancers as well as for earlier assessment of cancer recurrence and therapy efficacy. Despite their clinical interest, CTC analysis is not recommended by oncology guidelines so far. The main reason is that there is no gold standard technology for CTCs isolation and most of the current technologies are not yet validated for clinical use. In this chapter we will focus on the most relevant technologies for CTC isolation based on their properties and depending on whether it is a positive or negative selection. We also describe each technology based on its potential use and its relevance in breast cancer. The chapter also contains a future perspective including the challenges and requirements of CTC detection.


Assuntos
Neoplasias da Mama/patologia , Separação Celular/métodos , Células Neoplásicas Circulantes/patologia , Neoplasias da Mama/mortalidade , Contagem de Células , Humanos , Oncologia , Metástase Neoplásica/diagnóstico , Metástase Neoplásica/patologia , Recidiva Local de Neoplasia/diagnóstico , Recidiva Local de Neoplasia/patologia
20.
PLoS One ; 15(4): e0231473, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32315325

RESUMO

The aim of this study was to determine the effect of autologous serum (AS) eye drops on the density of human leucocyte antigen (HLA)-DR-positive epithelial cells and Langerhans cells on the ocular surface of patients with bilateral severe dry eye disease (DED) due to graft-versus-host disease (GvHD) or Sjögren's syndrome (SS). The study was conducted on 24 patients (48 eyes). AS was applied 6-10 times daily for 3 months together with regular artificial tear therapy. HLA-DR-positive cells were detected by direct immunocytochemistry on upper bulbar conjunctiva imprints obtained before and after treatment. The application of AS drops led to a statistically significant increase in the mean density of aberrant HLA-DR-positive conjunctival epithelial cells (p < 0.05) and HLA-DR-positive Langerhans cells (p < 0.05) in the GvHD group. Aberrant HLA-DR-positive epithelial cells in the SS group were decreased non-significantly. All patients reported a significant decrease in the Ocular Surface Disease Index (p < 0.01), which indicates improvement of the patient's subjective feelings after therapy. There was an expected but non-significant decrease of aberrant HLA-DR-positive conjunctival epithelial cells in the SS group only. However, the increased density of HLA-DR-positive cells, indicating slight subclinical inflammation, does not outweigh the positive effect of AS in patients with DED from GvHD.


Assuntos
Túnica Conjuntiva/metabolismo , Epitélio/metabolismo , Doença Enxerto-Hospedeiro/tratamento farmacológico , Doença Enxerto-Hospedeiro/metabolismo , Antígenos HLA-DR/metabolismo , Soro/metabolismo , Síndrome de Sjogren/metabolismo , Adulto , Idoso , Contagem de Células/métodos , Túnica Conjuntiva/efeitos dos fármacos , Síndromes do Olho Seco/tratamento farmacológico , Síndromes do Olho Seco/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Feminino , Humanos , Imuno-Histoquímica/métodos , Masculino , Pessoa de Meia-Idade , Soluções Oftálmicas/uso terapêutico , Síndrome de Sjogren/tratamento farmacológico
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