Phenotypic properties of transmitted founder HIV-1.

Defining the virus-host interactions responsible for HIV-1 transmission, including the phenotypic requirements of viruses capable of establishing de novo infections, could be important for AIDS vaccine development. Previous analyses have failed to identify phenotypic properties other than chemokine receptor 5 (CCR5) and CD4+ T-cell tropism that are preferentially associated with viral transmission. However, most of these studies were limited to examining envelope (Env) function in the context of pseudoviruses. Here, we generated infectious molecular clones of transmitted founder (TF; n = 27) and chronic control (CC; n = 14) viruses of subtypes B (n = 18) and C (n = 23) and compared their phenotypic properties in assays specifically designed to probe the earliest stages of HIV-1 infection. We found that TF virions were 1.7-fold more infectious (P = 0.049) and contained 1.9-fold more Env per particle (P = 0.048) compared with CC viruses. TF viruses were also captured by monocyte-derived dendritic cells 1.7-fold more efficiently (P = 0.035) and more readily transferred to CD4+ T cells (P = 0.025). In primary CD4+ T cells, TF and CC viruses replicated with comparable kinetics; however, when propagated in the presence of IFN-α, TF viruses replicated to higher titers than CC viruses. This difference was significant for subtype B (P = 0.000013) but not subtype C (P = 0.53) viruses, possibly reflecting demographic differences of the respective patient cohorts. Together, these data indicate that TF viruses are enriched for higher Env content, enhanced cell-free infectivity, improved dendritic cell interaction, and relative IFN-α resistance. These viral properties, which likely act in concert, should be considered in the development and testing of AIDS vaccines.

IL-22 mediates host defense against an intestinal intracellular parasite in the absence of IFN-γ at the cost of Th17-driven immunopathology.

The roles of Th1 and Th17 responses as mediators of host protection and pathology in the intestine are the subjects of intense research. In this study, we investigated a model of intestinal inflammation driven by the intracellular apicomplexan parasite Eimeria falciformis. Although IFN-γ was the predominant cytokine during E. falciformis infection in wild-type mice, it was found to be dispensable for host defense and the development of intestinal inflammation. E. falciformis-infected IFN-γR(-/-) and IFN-γ(-/-) mice developed dramatically exacerbated body weight loss and intestinal pathology, but they surprisingly harbored fewer parasites. This was associated with a striking increase in parasite-specific IL-17A and IL-22 production in the mesenteric lymph nodes and intestine. CD4(+) T cells were found to be the source of IL-17A and IL-22, which drove the recruitment of neutrophils and increased tissue expression of anti-microbial peptides (RegIIIß, RegIIIγ) and matrix metalloproteinase 9. Concurrent neutralization of IL-17A and IL-22 in E. falciformis-infected IFN-γR(-/-) mice resulted in a reduction in infection-induced body weight loss and inflammation and significantly increased parasite shedding. In contrast, neutralization of IL-22 alone was sufficient to increase parasite burden, but it had no effect on body weight loss. Treatment of an E. falciformis-infected intestinal epithelial cell line with IFN-γ, IL-17A, or IL-22 significantly reduced parasite development in vitro. Taken together, to our knowledge these data demonstrate for the first time an antiparasite effect of IL-22 during an intestinal infection, and they suggest that IL-17A and IL-22 have redundant roles in driving intestinal pathology in the absence of IFN-γ signaling.

The role of glycoprotein H of equine herpesviruses 1 and 4 (EHV-1 and EHV-4) in cellular host range and integrin binding.

Equine herpesvirus type 1 and 4 (EHV-1 and EHV-4) glycoprotein H (gH) has been hypothesized to play a role in direct fusion of the virus envelope with cellular membranes. To investigate gH's role in infection, an EHV-1 mutant lacking gH was created and the gH genes were exchanged between EHV-1 and EHV-4 to determine if gH affects cellular entry and/or host range. In addition, a serine-aspartic acid-isoleucine (SDI) integrin-binding motif present in EHV-1 gH was mutated as it was presumed important in cell entry mediated by binding to α4ß1 or α4ß7 integrins. We here document that gH is essential for EHV-1 replication, plays a role in cell-to-cell spread and significantly affects plaque size and growth kinetics. Moreover, we could show that α4ß1 and α4ß7 integrins are not essential for viral entry of EHV-1 and EHV-4, and that viral entry is not affected in equine cells when the integrins are inaccessible.

Comparison of proliferative and immunomodulatory potential of adipose-derived mesenchymal stem cells from young and geriatric cats.

Objectives The objective of this study was to compare the ability of adipose-derived mesenchymal stem cells (aMSCs) generated from young vs geriatric cats to proliferate in culture, suppress lymphocyte proliferation and undergo senescence. Methods Adipose tissues from five young (<5 years) and six geriatric (>10 years) cats were harvested and cryopreserved for subsequent aMSC isolation and culture. aMSC proliferation in culture was compared via determination of time until passage two and by 3-(4,5-demethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The immunomodulatory capacity of aMSCs was assessed using lymphocyte proliferation assays, and senescence was evaluated using senescence-associated B-galactosidase (SABG) expression. All assays were performed on aMSCs between passage two and passage three. Results aMSCs from geriatric cats took significantly longer ( P = 0.008) to reach passage two (median 11 days, range 9-22 days) compared with aMSCs from young healthy cats (median 7 days, range 6-8 days). No significant difference was detected between young and geriatric cats in terms of their ability to suppress lymphocyte proliferation. SABG expression was not significantly different between young and geriatric aMSCs. Conclusions and relevance Compared with young feline aMSCs, geriatric aMSCs are significantly impaired in their ability to rapidly proliferate to passage two following initial culture, presenting a concern for autologous therapy. Nonetheless, once the cells are expanded, young and geriatric cat aMSCs appear to be equivalent in terms of their ability to functionally suppress T-cell activation and proliferation.

Drug exposure and clinical effect of transdermal mirtazapine in healthy young cats: a pilot study.

Objectives The objective of this study was to measure drug exposure and clinical effects after administration of transdermal mirtazapine (TMZ) in healthy cats. Methods Phase I: seven healthy research cats received (1) 3.75 mg and 7.5 mg TMZ once aurally with 48 h serum sampling (serum samples were obtained via the jugular catheter at 0, 0.5, 1, 2, 5, 9, 12, 24, 36 and 48 h); (2) 7.5 mg TMZ and placebo daily aurally for 6 days then 48 h serum sampling; (3) 1.88 mg mirtazapine orally once with serum sampling at 1, 4 and 8 h. Phase II: 20 client-owned cats were enrolled in a randomized, double-blind, placebo-controlled, three-way crossover clinical effect study. Treatments consisted of 6 days of aural 7.5 mg TMZ or placebo gel at home, and 1.88 mg mirtazapine orally once in the clinic. Owners documented appetite, rate of food ingestion, begging activity and vocalization daily at home. On day 6, food consumed, activity and vocalization were documented in hospital, and trough and peak serum mirtazapine levels were obtained. Serum mirtazapine and gel concentrations were measured using liquid chromatography/tandem mass spectrometry. Results Phase I: administration of TMZ achieved measureable serum mirtazapine concentrations. Area under the curve0-48 of multidose 7.5 mg TMZ was significantly higher than single-dose 1.88 mg oral mirtazapine (OMZ) ( P = 0.02). Phase II: client-owned cats administered TMZ had a significant increase in appetite ( P = 0.003), rate of food ingestion ( P = 0.002), activity ( P = 0.002), begging ( P = 0.002) and vocalization ( P = 0.002) at home. In hospital there was a significant increase in food ingested with both TMZ and OMZ compared with placebo ( P <0.05). Gel concentrations ranged from 87%-119% of target dose. Conclusions and relevance TMZ 7.5 mg daily achieves measureable serum concentrations and produces significant appetite stimulation despite variance in compounded gel concentrations, but side effects denote a lower dose is indicated.

Assessment of absorption of transdermal ondansetron in normal research cats.

Objectives The objective of this study was to assess the absorption of transdermal ondansetron in healthy cats. Methods Five research cats with unremarkable complete blood count, biochemistry and urinalysis were used for both single- and multiple-dose application studies. For single-dose application, 4 mg ondansetron in 0.1 ml Lipoderm gel was applied once to the internal ear pinna. Blood samples were collected via jugular catheter over a 48 h period following administration (0, 15 mins, 30 mins, 1 h, 2 h, 4 h, 8 h, 12 h, 24 h and 48 h). For multiple-dose application, 4 mg ondansetron in 0.1 ml Lipoderm gel was applied for five consecutive days before blood samples were obtained in the same manner. Serum was separated and frozen prior to analysis. Ondansetron was measured via liquid chromatography coupled to tandem mass spectrometry. Results Analysis revealed no clinically relevant drug levels in serum after either single- or multiple-dose administration of 4 mg transdermal ondansetron. Conclusions and relevance Transdermal application of 4 mg ondansetron does not result in clinically relevant serum concentrations of drug. Despite characteristics of the drug that imply suitability for transdermal application, this does not appear to be an acceptable method of drug delivery for this medication at this dose. This study highlights the importance of assessing the suitability of each medication for transdermal administration.