Rotavirus antigen, cytokine, and neutralising antibody profiles in sera of children with and without HIV infection in Blantyre, Malawi

Background: Rotavirus and HIV infection are major causes of death among children in sub-Saharan Africa. A previous study reported no association between concomitant HIV infection and rotavirus disease severity among hospitalised children in Malawi. This study examined rotavirus antigenaemia and broader immune responses among HIV-infected and uninfected children.Methods: Stored (−80°C), paired sera from acute and convalescent phases of Malawian children less than 5 years old, hospitalised for acute gastroenteritis in the primary study, collected from July 1997 to June 1999, were utilised. Among children older than 15 months, HIV infection was defined as the presence of HIV antibody in the blood, when confirmed by at least 2 established methods. For those younger than 15 months, nested polymerase chain reaction (PCR) amplification of proviral DNA was used for verification. All were followed for up to 4 weeks after hospital discharge. Rotavirus antigen levels in sera were measured with Premier™ Rotaclone® rotavirus enzyme immunoassay (EIA) kit. Acute-phase sera were examined for 17 cytokines, using Luminex fluorescent bead human cytokine immunoassay kit. Rotavirus-specific IgA and neutralising activity were determined by EIA and microneutralisation (MN) assay, respectively. Human strains and bovine–human reassortants were propagated in MA104 cells with serum-free Iscove’s Modified Dulbecco’s Medium (IMDM). Differences in results, from specimens with and without HIV infection, were analysed for statistical significance using the chi-square test.Results: We detected rotavirus antigen in 30% of the HIV-infected and 21% HIV-uninfected, in the acute-phase sera. HIV-infected children developed slightly prolonged rotavirus antigenaemia compared to HIV-uninfected children.Conclusions: Rotavirus-specific IgA seroconversion rates and neutralising titres were similar in HIV-infected and HIV-uninfected children, thus, HIV infection had no major effect on immune responses to rotavirus infection

Puerarin Suppresses Invasion and Vascularization of Endometriosis Tissue Stimulated by 17β-Estradiol

BACKGROUND: Puerarin, a phytoestrogen with a weak estrogenic effect, binds to estrogen receptors, thereby competing with 17β-estradiol (E2) and producing an anti-estrogenic effect. This study was to investigate whether puerarin could suppress the invasion and vascularization of E2-stimulated endometriotic tissue. METHODOLOGY/PRINCIPAL FINDINGS: The endometriotic stromal cells (ESCs) were successfully established and their invasive ability under different treatments was assessed through a Transwell Assay. Simultaneously, matrix metallopeptidase 9 (MMP-9) and tissue inhibitor of metalloproteinase 1 (TIMP-1) were detected by western blotting. Vascularization of endometriotic tissues was observed by chicken chorioallantoic membrane (CAM) assay. The staining of MMP-9, intercellular adhesion molecule 1 (ICAM-1), TIMP-1, and vascular endothelial growth factor (VEGF) in grafted endometriotic tissues was examined using immunohistochemistry analysis. The purity of ESCs in isolated cells was >95%, as determined by the fluoroimmunoassay of vimentin. E2 (10(-8) mol/L) promoted the invasiveness of ESCs by increasing MMP-9 accumulation and decreasing TIMP-1 accumulation. Interestingly, puerarin (10(-9) mol/L) significantly reversed these effects (P<0.01). The CAM assay indicated that puerarin (10(-9) mol/L) also inhibited the angiopoiesis of endometriotic tissue stimulated by the E2 (10(-8) mol/L) treatment (P<0.05). Accordingly, immunohistochemistry showed that the accumulation of MMP-9, ICAM-1, and VEGF was reduced whereas that of TIMP-1 increased in the combination treatment group compared with the E2 treatment group. CONCLUSIONS/SIGNIFICANCE: This study demonstrated that puerarin could suppress the tissue invasion by ESCs and the vascularization of ectopic endometrial tissues stimulated by E2, suggesting that puerarin may be a potential drug for the treatment of endometriosis

Distribution, Quantification and Toxicity of Cinnamaldehyde in Electronic Cigarette Refill Fluids and Aerosols

Objective The aim of this study was to evaluate the distribution, concentration and toxicity of cinnamaldehyde in electronic cigarette (e-cigarette) refill fluids and aerosols. Methods The distribution and concentration of cinnamaldehyde were determined in 39 e-cigarette refill fluids plus 6 duplicates using gas chromatography and mass spectrometry (GC/MS). A cinnamaldehyde toxicity profile was established for embryonic and adult cells using a live cell imaging assay, immunocytochemistry, the comet assay and a recovery assay. Results Twenty of the 39 refill fluids contained cinnamaldehyde at concentrations that are cytotoxic to human embryonic and lung cells in the MTT assay. Cinnamon Ceylon aerosol produced in a cartomizer-style e-cigarette was cytotoxic. Cinnamon Ceylon aerosols and refill fluid aerosols (80% propylene glycol or cinnamaldehyde/propylene glycol) made using a tank/boxmod e-cigarette were more cytotoxic at 5 V than 3 V. Using GC/MS, aerosols produced at 5 V contained 10 additional peaks not present in aerosol generated at 3 V. One of these, 2,3-butandione (diacetyl), was confirmed with an authentic standard. Cinnamaldehyde depolymerised microtubules in human pulmonary fibroblasts. At concentrations that produced no effect in the MTT assay, cinnamaldehyde decreased growth, attachment and spreading; altered cell morphology and motility; increased DNA strand breaks; and increased cell death. At the MTT IC50 concentration, lung cells were unable to recover from cinnamaldehyde after 2 hours of treatment, whereas embryonic cells recovered after 8 hours. Conclusions Cinnamaldehyde-containing refill fluids and aerosols are cytotoxic, genotoxic and low concentrations adversely affect cell processes and survival. These data indicate that cinnamaldehyde in e-cigarette refill fluids/aerosols may impair homeostasis in the respiratory system

Role for 18:1 lysophosphatidic acid as an autocrine mediator in prostate cancer cells

Lysophosphatidic acid (LPA) is a lipid mediator that may play an important role in growth and survival of carcinomas. In this study, LPA production and response were characterized in two human prostate cancer (CaP) cell lines: PC-3 and Du145. Bombesin, a neuroendocrine peptide that is mitogenic for CaP cells, stimulated focal adhesion kinase phosphorylation and activated the extracellular signal-regulated kinase/mitogen-activated protein kinase pathway. Similar responses were elicited by 18:1 LPA (oleoyl-LPA). Studies using radioisotopic labeling revealed that both PC-3 and Du145 generate LPA and that LPA production is increased by bombesin. The kinetics of bombesin-induced phospholipase D activation and LPA production were similar. Using electrospray ionization mass spectrometry, 18:1 LPA was found to be an abundant LPA species in CaP cell medium. Structure activity studies of acyl-LPAs revealed that 18:1 LPA is most efficacious for activation of extracellular signal-regulated kinase and phospholipase D in CaP cells. Incubation with 18:1 LPA caused homologous desensitization of LPA response, whereas bombesin caused heterologous desensitization. LPA was present at nanomolar levels in medium from bombesin-treated cells. LPA extracted from the medium induced calcium mobilization in CaP cells. These results demonstrate that bioactive LPA is generated by CaP cells in response to a mitogen and suggest that 18:1 LPA can act as an autocrine mediator

Pro-and Anti-inflammatory Forms of Interleukin-1 in the Tear Fluid and Conjunctiva of Patients with Dry-Eye Disease

PURPOSE. To compare the expression of the pro-and antiinflammatory forms of interleukin (IL)-1 in the tear fluid and conjunctival epithelium of normal eyes and those with dry-eye disease. METHODS. The concentrations of IL-1␣, IL-1␤ (precursor and mature forms), and IL-1 receptor antagonist (IL-1Ra) were measured by ELISA in tear fluid samples obtained from normal individuals and patients with dry eye who had rosacea-associated meibomian gland disease (MGD) or Sjögren&apos;s syndrome (SS) aqueous tear deficiency (ATD). These cytokines were also measured in normal tear fluid before and after nasal stimulation to induce reflex tearing. The relative expression of these cytokines was evaluated in conjunctival impression cytology specimens and conjunctival biopsy tissue obtained from normal subjects and SS ATD-affected patients using immunofluorescent staining. Matrix metalloproteinase (MMP)-9 concentration and activity in the tear fluid were evaluated with gelatin zymography and with an MMP-9 activity assay kit, respectively. RESULTS. Compared with normal subjects, the concentration of IL-1␣ and mature IL-1␤ in the tear fluid was increased, and the concentration of precursor IL-1␤ was decreased in patients with MGD (P Ͻ 0.05, P ϭ 0.02, and P Ͻ 0.01, respectively) and SS ATD (P Ͻ 0.001, P ϭ 0.02, and P Ͻ 0.001, respectively). There was no significant change in the concentration of IL-1␣, precursor IL-1␤, and IL-1Ra in reflex tear fluid, indicating that the lacrimal glands may secrete these cytokines. The activity of MMP-9, a physiological activator of IL-1␤, was significantly elevated in the tear fluid of both dry-eye groups compared with normal subjects. A strong positive correlation was observed between the intensity of corneal fluorescein staining and the tear fluid IL-1␣ concentration (r 2 ϭ 0.17, P Ͻ 0.02) and the mature-to-precursor IL-1␤ ratio (r 2 ϭ 0.46, P Ͻ 0.001). Positive immunofluorescent staining for IL-1␣, mature IL-1␤, and IL-1Ra was observed in a significantly greater percentage of conjunctival cytology specimens from eyes with SS ATD than in those from normal eyes (P Ͻ 0.01 for IL-1␣, P Ͻ 0.009 for mature IL-1␤, and P Ͻ 0.05 for IL-1Ra)

Menthol in Electronic Cigarettes: A Contributor to Respiratory Disease?

Menthol is widely used in tobacco products. This study compared the effects of menthol on human bronchial epithelium using submerged cultures, a VITROCELL® cloud chamber that provides air liquid interface (ALI) exposure without solvents or heating, and a Cultex ALI system that delivers aerosol equivalent to that inhaled during vaping. In submerged culture, menthol significantly increased calcium influx and mitochondrial reactive oxygen species (ROS) via the TRPM8 receptor, responses that were inhibited by a TRPM8 antagonist. VITROCELL® cloud chamber exposure of BEAS-2B monolayers increased mitochondrial protein oxidation, expression of the antioxidant enzyme SOD2, activation of NF-κB, and secretion of inflammatory cytokines (IL-6 and IL-8). Proteomics data collected following ALI exposure of 3D EpiAirway tissue in the Cultex showed upregulation of NRF-2-mediated oxidative stress, oxidative phosphorylation, and IL-8 signaling. Across the three platforms, menthol adversely effected human bronchial epithelium in a manner that could lead to respiratory disease