Impaired vascular responses to parasympathetic nerve stimulation and muscarinic receptor activation in the submandibular gland in non-obese diabetic mice

Introduction Decreased vascular responses to salivary gland stimulation are observed in Sjögren's syndrome patients. We investigate whether impaired vascular responses to parasympathetic stimulation and muscarinic receptor activation in salivary glands parallels hyposalivation in an experimental model for Sjögren's syndrome. Methods Blood flow responses in the salivary glands were measured by laser Doppler flowmeter. Muscarinic receptor activation was followed by saliva secretion measurements. Nitric oxide synthesis-mediated blood flow responses were studied after administration of a nitric oxide synthase inhibitor. Glandular autonomic nerves and muscarinic 3 receptor distributions were also investigated. Results Maximal blood flow responses to parasympathetic stimulation and muscarinic receptor activation were significantly lower in nonobese diabetic (NOD) mice compared with BALB/ c mice, coinciding with impaired saliva secretion in nonobese diabetic mice (P < 0.005). Nitric oxide synthase inhibitor had less effect on blood flow responses after parasympathetic nerve stimulation in nonobese diabetic mice compared with BALB/c mice (P < 0.02). In nonobese diabetic mice, salivary gland parasympathetic nerve fibres were absent in areas of focal infiltrates. Muscarinic 3 receptor might be localized in the blood vessel walls of salivary glands. Conclusions Impaired vasodilatation in response to parasympathetic nerve stimulation and muscarinic receptor activation may contribute to hyposalivation observed in nonobese diabetic mice. Reduced nitric oxide signalling after parasympathetic nerve stimulation may contribute in part to the impaired blood flow responses. The possibility of muscarinic 3 receptor in the vasculature supports the notion that muscarinic 3 receptor autoantibodies present in nonobese diabetic mice might impair the fluid transport required for salivation. Parasympathetic nerves were absent in areas of focal infiltrates, whereas a normal distribution was found within glandular epithelium.publishedVersio

Effect of D222G Mutation in the Hemagglutinin Protein on Receptor Binding, Pathogenesis and Transmissibility of the 2009 Pandemic H1N1 Influenza Virus

Influenza viruses isolated during the 2009 H1N1 pandemic generally lack known molecular determinants of virulence associated with previous pandemic and highly pathogenic avian influenza viruses. The frequency of the amino acid substitution D222G in the hemagglutinin (HA) of 2009 H1N1 viruses isolated from severe but not mild human cases represents the first molecular marker associated with enhanced disease. To assess the relative contribution of this substitution in virus pathogenesis, transmission, and tropism, we introduced D222G by reverse genetics in the wild-type HA of the 2009 H1N1 virus, A/California/04/09 (CA/04). A dose-dependent glycan array analysis with the D222G virus showed a modest reduction in the binding avidity to human-like (α2-6 sialylated glycan) receptors and an increase in the binding to avian-like (α2-3 sialylated glycan) receptors in comparison with wild-type virus. In the ferret pathogenesis model, the D222G mutant virus was found to be similar to wild-type CA/04 virus with respect to lethargy, weight loss and replication efficiency in the upper and lower respiratory tract. Moreover, based on viral detection, the respiratory droplet transmission properties of these two viruses were found to be similar. The D222G virus failed to productively infect mice inoculated by the ocular route, but exhibited greater viral replication and weight loss than wild-type CA/04 virus in mice inoculated by the intranasal route. In a more relevant human cell model, D222G virus replicated with delayed kinetics compared with wild-type virus but to higher titer in human bronchial epithelial cells. These findings suggest that although the D222G mutation does not influence virus transmission, it may be considered a molecular marker for enhanced replication in certain cell types.Centers for Disease Control and Prevention (U.S.)United States. National Institutes of Health (merit award R37 GM057073-13)Singapore-MIT Alliance for Research and Technolog

RISK ASSESSMENT IN CORRELATION WITH OBESITY AND LABORATORY MONITORING OF THE MOST COMMON FEMALE BREAST CANCERS IN SARAJEVO CANTON

Introduction: Breast cancer (BC) is a malignant disease that predominantly affects women, with known genetic components such as mutations in tumor suppressor genes BRCA1 and BRCA2. Other risk factors include unhealthy lifestyles, lack of physical activity, and consumption of alcohol and cigarettes. Aging also plays a role in BC development, with hormonal influences such as estrogen and progesterone promoting cancer growth. Material and Methods: Research was conducted using data collection tools for risk factors and tumor markers from primary healthcare unit records. The sample comprised 200 women, divided into two groups based on BC diagnosis, with complete medical documentation. Male BC cases were excluded. Results: Statistical significance was found between genetic components, family history, aging, obesity, alcohol and cigarette consumption, longer hormone exposure, and female BC development using the Chi-Square test, confirmed by Fisher’s Exact test. Tumor markers CA 15-3, CEA, CA 19-9, and CA 125 were useful for BC screening and metastasis detection, as determined by the One Sample T-test. In Sarajevo Canton, invasive ductal BC was the most common type among women, while lobular carcinoma in situ was the least common. Conclusion: Correlations between risk factors, including aging, unhealthy lifestyles, and hormone exposure, and increased BC risk were confirmed. Tumor markers CA 15-3, CEA, CA 19-9, and CA 125 were effective in diagnosis, screening, and metastasis detection in females, with sensitivity for regression detection at 81.8% and specificity at 100%

DIFFERENCES BETWEEN BIOCHEMICAL, HEMATOLOGICAL AND COAGULATION PARAMETERS AMONG PATIENTS WITH MILD AND SEVERE COVID-19

Introduction: COVID-19 is an infectious disease caused by the severe acute respiratory syndrome coronavirus 2 and causes a series of respiratory symptoms. Considering the appearance and development of symptoms, the course of COVID-19 can go from mild to severe. Depending on the course of COVID-19, the laboratory parameters change a lot, trying to defend the organism against the foreign pathogen and all the changes it causes.Therefore, the aim of this study is to observe the differences between biochemical, hematological and coagulation parameters depending on the disease stage of COVID-19 patients. Material and methods: We conducted a cross-sectional study which included 160 COVID-19 patients from Sarajevo, Bosnia and Herzegovina. Biochemical, hematological and coagulation analyses were performed. Results: COVID-19 patients with a severe clinical course have higher average values of fibrinogen (6.53±4.47,p<0.001), D-dimer (6.89±7.81, p<0.001), APTT (32.05±5.96, p=0.002), eosinophil (0.66±0.09, p=0.002) and CRP (93.42±75.86, p= 0.023), and lower values of lymphocytes (1.04±0.98, p<0.001), monocytes (0.45±0.3, p<0.001), compared to COVID-19 patients with a mild clinical course. COVID-19 patients with a severe clinical course had higher average values of neutrophils (10.12±5.80, p=0.002) and lower values of reactive lymphocytes (0.02±0.03, p<0.001) compared to COVID-19 patients with a mild clinical course. Conclusion: Biochemical, hematological and coagulation parameters can be a sensitive and specific biomarker for distinction of mild and severe COVID-19

Sialic acid receptor detection in the human respiratory tract: evidence for widespread distribution of potential binding sites for human and avian influenza viruses

<p>Abstract</p> <p>Background</p> <p>Influenza virus binds to cell receptors via sialic acid (SA) linked glycoproteins. They recognize SA on host cells through their haemagglutinins (H). The distribution of SA on cell surfaces is one determinant of host tropism and understanding its expression on human cells and tissues is important for understanding influenza pathogenesis. The objective of this study therefore was to optimize the detection of α2,3-linked and α2,6-linked SA by lectin histochemistry by investigating the binding of Sambucus nigra agglutinin (SNA) for SAα2,6Gal and Maackia amurensis agglutinin (MAA) for SAα2,3Gal in the respiratory tract of normal adults and children.</p> <p>Methods</p> <p>We used fluorescent and biotinylated SNA and MAA from different suppliers on archived and prospectively collected biopsy and autopsy specimens from the nasopharynx, trachea, bronchus and lungs of fetuses, infants and adults. We compared different methods of unmasking for tissue sections to determine if these would affect lectin binding. Using serial sections we then compared the lectin binding of MAA from different suppliers.</p> <p>Results</p> <p>We found that unmasking using microwave treatment in citrate buffer produced increased lectin binding to the ciliated and glandular epithelium of the respiratory tract. In addition we found that there were differences in tissue distribution of the α2,3 linked SA when 2 different isoforms of MAA (MAA1 and MAA2) lectin were used. MAA1 had widespread binding throughout the upper and lower respiratory tract and showed more binding to the respiratory epithelium of children than in adults. By comparison, MAA2 binding was mainly restricted to the alveolar epithelial cells of the lung with weak binding to goblet cells. SNA binding was detected in bronchial and alveolar epithelial cells and binding of this lectin was stronger to the paediatric epithelium compared to adult epithelium. Furthermore, the MAA lectins from 2 suppliers (Roche and EY Labs) tended to only bind in a pattern similar to MAA1 (Vector Labs) and produced a different binding pattern to MAA2 from Vector Labs.</p> <p>Conclusion</p> <p>The lectin binding pattern of MAA may vary depending on the supplier and the different isoforms of MAA show a different tissue distribution in the respiratory tract. This finding is important if conclusions about the potential binding sites of SAα2,3 binding viruses, such as influenza or human parainfluenza are to be made.</p

Apoptosis and Pathogenesis of Avian Influenza A (H5N1) Virus in Humans

Apoptosis may play a crucial role in the pathogenesis of pneumonia and lymphopenia caused by this virus in humans

The mouse model is suitable for the study of viral factors governing transmission and pathogenesis of highly pathogenic avian influenza (HPAI) viruses in mammals

Highly pathogenic avian influenza (HPAI) viruses of the H5 and H7 subtype pose a major public health threat due to their capacity to cross the species barrier and infect mammals, for example dogs, cats and humans. In the present study we tested the capacity of selected H7 and H5 HPAI viruses to infect and to be transmitted from infected BALB/c mice to contact sentinels. Previous experiments have shown that viruses belonging to both H5 and H7 subtypes replicate in the respiratory tract and central nervous system of experimentally infected mice. In this study we show that selected H7N1 and H5N1 HPAI viruses can be transmitted from mouse-to-mouse by direct contact, and that in experimentally infected animals they exhibit a different pattern of replication and transmission. Our results can be considered as a starting point for transmission experiments involving other influenza A viruses with α 2-3 receptor affinity in order to better understand the viral factors influencing transmissibility of these viruses in selected mammalian species

Guinea Pig Model for Evaluating the Potential Public Health Risk of Swine and Avian Influenza Viruses

BACKGROUND: The influenza viruses circulating in animals sporadically transmit to humans and pose pandemic threats. Animal models to evaluate the potential public health risk potential of these viruses are needed. METHODOLOGY/PRINCIPAL FINDINGS: We investigated the guinea pig as a mammalian model for the study of the replication and transmission characteristics of selected swine H1N1, H1N2, H3N2 and avian H9N2 influenza viruses, compared to those of pandemic (H1N1) 2009 and seasonal human H1N1, H3N2 influenza viruses. The swine and avian influenza viruses investigated were restricted to the respiratory system of guinea pigs and shed at high titers in nasal tracts without prior adaptation, similar to human strains. None of the swine and avian influenza viruses showed transmissibility among guinea pigs; in contrast, pandemic (H1N1) 2009 virus transmitted from infected guinea pigs to all animals and seasonal human influenza viruses could also horizontally transmit in guinea pigs. The analysis of the receptor distribution in the guinea pig respiratory tissues by lectin histochemistry indicated that both SAα2,3-Gal and SAα2,6-Gal receptors widely presented in the nasal tract and the trachea, while SAα2,3-Gal receptor was the main receptor in the lung. CONCLUSIONS/SIGNIFICANCE: We propose that the guinea pig could serve as a useful mammalian model to evaluate the potential public health threat of swine and avian influenza viruses

Ciliary Beating Recovery in Deficient Human Airway Epithelial Cells after Lentivirus Ex Vivo Gene Therapy

Primary Ciliary Dyskinesia is a heterogeneous genetic disease that is characterized by cilia dysfunction of the epithelial cells lining the respiratory tracts, resulting in recurrent respiratory tract infections. Despite lifelong physiological therapy and antibiotics, the lungs of affected patients are progressively destroyed, leading to respiratory insufficiency. Recessive mutations in Dynein Axonemal Intermediate chain type 1 (DNAI1) gene have been described in 10% of cases of Primary Ciliary Dyskinesia. Our goal was to restore normal ciliary beating in DNAI1–deficient human airway epithelial cells. A lentiviral vector based on Simian Immunodeficiency Virus pseudotyped with Vesicular Stomatitis Virus Glycoprotein was used to transduce cultured human airway epithelial cells with a cDNA of DNAI1 driven by the Elongation Factor 1 promoter. Transcription and translation of the transduced gene were tested by RT–PCR and western blot, respectively. Human airway epithelial cells that were DNAI1–deficient due to compound heterozygous mutations, and consequently had immotile cilia and no outer dynein arm, were transduced by the lentivirus. Cilia beating was recorded and electron microscopy of the cilia was performed. Transcription and translation of the transduced DNAI1 gene were detected in human cells treated with the lentivirus. In addition, immotile cilia recovered a normal beat and outer dynein arms reappeared. We demonstrated that it is possible to obtain a normalization of ciliary beat frequency of deficient human airway epithelial cells by using a lentivirus to transduce cells with the therapeutic gene. This preliminary step constitutes a conceptual proof that is indispensable in the perspective of Primary Ciliary Dyskinesia's in vivo gene therapy. This is the first time that recovery of cilia beating is demonstrated in this disease