RESUMO
The conducting airways are lined by distinct cell types, comprising basal, secretory, ciliated, and rare cells, including ionocytes, solitary cholinergic chemosensory cells, and solitary and clustered (neuroepithelial bodies) neuroendocrine cells. Airway neuroendocrine cells are in clinical focus since they can give rise to small cell lung cancer. They have been implicated in diverse functions including mechanosensation, chemosensation, and regeneration, and were recently identified as regulators of type 2 immune responses via the release of the neuropeptide calcitonin gene-related peptide (CGRP). We here assessed the expression of the chemokine CXCL13 (B cell attracting chemokine) by these cells by RT-PCR, in silico analysis of publicly available sequencing data sets, immunohistochemistry, and immuno-electron microscopy. We identify a phenotype of neuroendocrine cells in the naïve mouse, producing the chemokine CXCL13 predominantly in solitary neuroendocrine cells of the tracheal epithelium (approx. 70% CXCL13+) and, to a lesser extent, in the solitary neuroendocrine cells and neuroepithelial bodies of the intrapulmonary bronchial epithelium (< 10% CXCL13+). In silico analysis of published sequencing data of murine tracheal epithelial cells was consistent with the results obtained by immunohistochemistry as it revealed that neuroendocrine cells are the major source of Cxcl13-mRNA, which was expressed by 68-79% of neuroendocrine cells. An unbiased scRNA-seq data analysis of overall gene expression did not yield subclusters of neuroendocrine cells. Our observation demonstrates phenotypic heterogeneity of airway neuroendocrine cells and points towards a putative immunoregulatory role of these cells in bronchial-associated lymphoid tissue formation and B cell homeostasis.
Assuntos
Quimiocina CXCL13 , Células Neuroendócrinas , Animais , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Colinérgicos , Células Epiteliais/metabolismo , Pulmão/metabolismo , Camundongos , Células Neuroendócrinas/metabolismo , RNA Mensageiro/genética , TraqueiaRESUMO
Disorders of gallbladder motility can lead to serious pathology. Bitter tastants acting upon bitter taste receptors (TAS2R family) have been proposed as a novel class of smooth muscle relaxants to combat excessive contraction in the airways and other organs. To explore whether this might also emerge as an option for gallbladder diseases, we here tested bitter tastants for relaxant properties and profiled Tas2r expression in the mouse gallbladder. In organ bath experiments, the bitter tastants denatonium, quinine, dextromethorphan, and noscapine, dose-dependently relaxed the pre-contracted gallbladder. Utilizing gene-deficient mouse strains, neither transient receptor potential family member 5 (TRPM5), nor the Tas2r143/Tas2r135/Tas2r126 gene cluster, nor tuft cells proved to be required for this relaxation, indicating direct action upon smooth muscle cells (SMC). Accordingly, denatonium, quinine and dextromethorphan increased intracellular calcium concentration preferentially in isolated gallbladder SMC and, again, this effect was independent of TRPM5. RT-PCR revealed transcripts of Tas2r108, Tas2r126, Tas2r135, Tas2r137, and Tas2r143, and analysis of gallbladders from mice lacking tuft cells revealed preferential expression of Tas2r108 and Tas2r137 in tuft cells. A TAS2R143-mCherry reporter mouse labeled tuft cells in the gallbladder epithelium. An in silico analysis of a scRNA sequencing data set revealed Tas2r expression in only few cells of different identity, and from in situ hybridization histochemistry, which did not label distinct cells. Our findings demonstrate profound tuft cell- and TRPM5-independent relaxing effects of bitter tastants on gallbladder smooth muscle, but do not support the concept that these effects are mediated by bitter receptors.
Assuntos
Vesícula Biliar , Músculo Liso , Receptores Acoplados a Proteínas G , Canais de Cátion TRPM , Animais , Camundongos , Cálcio/metabolismo , Dextrometorfano/farmacologia , Vesícula Biliar/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Relaxamento Muscular/efeitos dos fármacos , Músculo Liso/metabolismo , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , Noscapina/farmacologia , Compostos de Amônio Quaternário/farmacologia , Quinina/farmacologia , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Transdução de Sinais , Paladar/fisiologia , Canais de Cátion TRPM/metabolismo , Canais de Cátion TRPM/genética , Células em Tufo/metabolismoRESUMO
Cyclophosphamide (CP) is a widely used anticancer and immunosuppressant that induces oxidative stress. To ameliorate the side effects resulted from CP treatment, liposomes were tested as an efficient drug delivery system with or without vitamin C as an antioxidant. CP resulted in clastogenic and cytotoxic effects that significantly increased for the total chromosomal aberrations as well as the numerical ones in the CP group (150.8 and 6, respectively) than the control group (6.6 and 0.0) as mean values at p < 0.05. Micronucleus assay showed a significant increased micronucleated polychromatic erythrocytes percentage (MNPCEs% = 11.7%) and a significant decrease of polychromatic to normochromatic erythrocytes ratio (0.551) when compared to the group treated with liposomised CP and vitamin C (3.44%; 0.795, respectively) at p < 0.05. Also, the total glutathione S-transferase activity as a body antioxidant enzyme was decreased from 52.2 in the control to 16.09 nmol/min/mg protein in CP group at p < 0.05, while the highly significant amelioration results were observed in the liposomised vitamin C and CP group (40.88 nmol/min/mg protein). Our findings support the potential use of CP in a liposomal formulation doped with vitamin C to diminish the potential side effects of the agent.
Assuntos
Ácido Ascórbico/farmacologia , Cápsulas/química , Aberrações Cromossômicas/induzido quimicamente , Ciclofosfamida/toxicidade , Lipossomos/química , Animais , Antineoplásicos Alquilantes/química , Antineoplásicos Alquilantes/toxicidade , Ácido Ascórbico/química , Ciclofosfamida/química , Relação Dose-Resposta a Droga , Masculino , CamundongosRESUMO
BACKGROUND: Radiographs have been used for forensic identification purpose. At times when only skull remains are found and other means of identification fail, radiographs of skull may be used for identification purpose. AIM: The objective of this study was morphometric evaluation of the frontal sinus by using digital posteroanterior skull radiograph in relation to age and gender and to establish its forensic importance. MATERIALS AND METHODS: The study was conducted at Alfarabi Private College for Dentistry and Nursing, Jeddah, Kingdom of Saudi Arabia. It included 400 subjects (200 males and 200 females), aged 14-70 years. Radiographs of the individuals were taken by digital radiography, and morphometric evaluation of frontal sinus was carried out by using Adobe Photoshop CS3 Extended. RESULTS: Unilateral absence of sinus was noted in 2.5% males and 1.5% females. Bilateral absence was noticed in 2% males and 0.5% females. Right and left frontal sinus symmetry was seen in 83.20% of the individuals. The left-dominated asymmetry was observed in 6.98% individuals. The right-dominated asymmetry was observed in 9.82% individuals. Simple logistic regression analysis of gender by different variables showed right width and left width, which are most suited regressors for sex determination. The rate of accuracy in classification of males and females varied from 67.70% to 95.90%. Stepwise multiple regression analysis of estimation of age by different variables showed right sinus height is the best predictor of age. CONCLUSION: In this study, the radiographic images of the frontal sinus showed significant morphological difference in relation to age and gender in Saudi population. On the basis of this evidence, it is proposed that the morphologic evaluation of frontal sinus can be used for personal identification.
RESUMO
AIM: The main objectives of this study were to determine the prevalence of fascioliasis infections in cattle and buffaloes, slaughtered in El-Kharga city slaughterhouse at New Valley Governorate. MATERIALS AND METHODS: The slaughtered animals were daily inspected for liver fascioliasis allover 2016. Macroscopic fascioliasis was detected from a total of 2251 basing on animals specie, sex, season, and Fasciola spp. in addition to microscopic examination of blood, fecal samples which collected from female cattle and buffalo (50 each). RESULTS: The total prevalence rate of Fasciola sp. infection occurs in the study area were about 695/2251 (30.88%) from the total cattle and bovine slaughtered carcasses. The incidence of fascioliasis was 4/12 (33.33%) and 678/2200 (30.82%) for females and males cattle carcasses, respectively, while the infection rate in buffalo carcasses was 1/4 (25.00%) and 12/35 (34.29%) for females and males buffalo carcasses, respectively. CONCLUSION: The moderate fasciolosis infection in cattle and buffaloes slaughtered at the municipal abattoir of El-Kharga, Egypt. The highest fascioliasis infection was recorded during winter and autumn. It constitutes a major cause of economic losses at El-Kharga abattoir and threat public health.
RESUMO
The utility of a new instrument for rapid virus quantitation, the Virus Counter, was evaluated in a blind study conducted at three sites. This instrument is a substantially improved version of the original academic research instrument described previously by Stoffel and Rowlen (2005a). The addition of hydrodynamic focusing, a self-contained fluidics system and customized software for system control and data analysis has resulted in a commercially viable and available design. Baculovirus samples were provided by Protein Sciences Corporation and blinded to InDevR and Baylor College of Medicine. Protein Sciences Corporation and Baylor College of Medicine analyzed the samples by plaque assay and InDevR analyzed the samples using the Virus Counter. Serial dilution of stock viruses into growth media and buffer allowed for comparison of measured versus intended concentrations. Direct log-scale comparison between pooled Virus Counter results and pooled plaque assay results indicated a linear relationship (slope=1.1±0.2, R(2)=0.86) with statistically significant Pearson correlation (r=0.93, p<0.001).
Assuntos
Baculoviridae/isolamento & purificação , Carga Viral , Ensaio de Placa ViralRESUMO
A robust and reliable GMP-compatible fed-batch process was successfully developed for the production of recombinant hemagglutinin (rHA) proteins by expresSF cells. The feeding solution, feeding strategy as well as the cell density at infection were optimized to maximize the final rHA production yields without affecting the existing rHA recovery protocol and downstream process. A simple and stable feeding solution was formulated and a rational feeding regimen designed to yield, depending on the rHA baculovirus used, between 2- and 3-fold enhancements in volumetric rHA production with increased specific productivity compared to the batch culture. Recombinant HA from fed-batch cultures could be simply recovered following cell lysis and purified through chromatographic steps. Overall, the increased rHA yield was maintained throughout the whole process. The performance, reproducibility and scalability of the fed-batch process was successfully demonstrated in 12 bioreactor runs of 2- and 10-L working volume using five different rHA encoding baculoviruses.
Assuntos
Vacinas contra Influenza/biossíntese , Vacinas contra Influenza/metabolismo , Engenharia de Proteínas/métodos , Proteínas Recombinantes/imunologia , Animais , Linhagem Celular , Hemaglutininas/genética , Hemaglutininas/metabolismo , Vacinas contra Influenza/genética , Vacinas contra Influenza/imunologia , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , SpodopteraRESUMO
Numerous human infections with avian influenza viruses in Asia in recent years have raised the concern that the next influenza pandemic is imminent. The most effective way to combat influenza is through the vaccination of the public. However, a minimum of 3-6 months is needed to develop an influenza vaccine using the traditional egg-based vaccine approach. The influenza hemagglutinin protein (HA), the active ingredient in the current vaccine, can be expressed in insect cells using the baculovirus expression vector system and purified rapidly. An influenza vaccine based on such a recombinant antigen allows a more timely response to a potential influenza pandemic. Here, we report an innovative monitoring assay for recombinant HA (rHA) expression and a rapid purification process. Various biochemical analyses indicate that the purified rHA is properly folded and biologically active.