Clinical laboratory indicators in study subjects with PUUV nephropathia epidemica.

a<p>AKI = acute kidney injury; severe AKI = >3-fold rise in serum creatinine during acute illness compared to a 6 month baseline value.</p>b<p>Values are median (95% confidence interval of median).</p

Urinary sediment GATA-3 mRNA levels are elevated and T-bet mRNA levels are decreased during acute illness in PUUV-induced severe AKI.

<p>Severe AKI is >3-fold rise in serum Cr during acute illness compared to a 6 month baseline. Illness day 1 is the first calendar day of reported fever. a) Urinary sediment CD3ε mRNA relative expression compared to 2 week baseline value b) Urinary sediment GATA-3 mRNA relative expression compared to 2 week baseline value c) Urinary sediment T-bet mRNA relative expression compared to 2 week baseline value. Symbols and error bars are mean±S.E.</p

Multivariate statistical models for acute kidney injury in study subjects with PUUV nephropathia epidemica.

a<p>Odds ratio (OR) for developing severe PUUV-induced acute kidney injury (AKI) compared to non-severe AKI. Severe AKI = >3-fold rise in serum creatinine during acute illness compared to a 6 month baseline value.</p>b<p>95% confidence interval (CI) for the Odds Ratio.</p>c<p>log₁₀ transformed variable.</p>d<p>White blood cell (WBC).</p>e<p>Linear regression of log₁₀ transformed peak creatinine ratio vs. log₁₀ transformed variables.</p>f<p>95% confidence interval (CI) for the linear regression coefficient.</p

Characteristics of study subjects with PUUV nephropathia epidemica.

a<p>AKI = acute kidney injury; severe AKI = >3-fold rise in serum creatinine during acute illness compared to a 6 month baseline value.</p>b<p>Values are median (95% confidence interval of median).</p

Maximum urine IL-6, plasma IL-6, and urine IL-8 concentrations are positively correlated with the degree of PUUV-induced AKI.

<p>a) maximum urine IL-6 levels b) maximum plasma IL-6 levels c) maximum urine IL-8 levels d) maximum plasma IL-8 levels.</p

Weight loss curves of individual mice and splenocyte counts following i.n. VACV infection of C57/BL6 mice.

<p>(A) Groups of female C57/BL6 mice (n = 5–8) were infected with 10⁵, 10⁶ and 10⁷ PFU of vGK5 by the i.n. route. The percentage of weight relative to the initial body weight (100%) was plotted and the data are presented as percent change in body weight following infection. † depicts days that individual mice were last alive. (B) Average spleen counts±standard deviation of mice were assessed by trypan blue exclusion at days 3, 5 and 7 post infection with 10⁶ VACV-WR and vGK5 by the i.n. route. Data shown are representative of 2 experiments performed and demonstrate that high dose VACV-WR i.n. infections result in significantly (p<0.05) lower lymphocytes in the spleens during acute infection.</p

Viral titers following i.n. infections.

<p>(A) Spleens and (B) lungs (n = 4–8 per group) were harvested on day 7 from mice infected with VACV-WR and vGK5 by the i.n. route. VACV titers were determined and expressed as log₁₀ PFU per gram of lung and spleen tissue. – represents median values of titers in respective organs. N.S. = not significant. Each symbol represents the titer obtained in target organs of individual mice; median values are denoted by horizontal lines. P values were determined by Student's <i>t</i> test.</p

Immune responses following infection by the tail scarification and i.p. routes.

<p>Mice were infected with 1×10⁶ PFU VACV-WR or vGK5 by the i.p. and t.s. routes. (A) Lung lymphocytes and splenocytes obtained from mice (n = 4 mice/group except for infection with VACV-WR by the t.s. route where splenocytes and lung lymphocytes from 2 mice were pooled together) infected 7 days prior were stained with B8R_20–27 tetramer. The data shown represent frequencies of cells that were tetramer positive within the CD3+CD8+ gate. Each symbol represents the frequency of tetramer+ T cells obtained in target organs of individual mice; median values are denoted by horizontal lines. (B) Seven days post infection, splenocytes were isolated and CTL assays were carried out using RMA cells infected with VACV-WR (moi = 5), vGK5 (moi = 5) at different (E/T) ratios. Data shown are representative of 2–3 experiments performed for each condition for the i.p. route. (C) PRNT50 antibody titers were measured in sera of mice immunized 3 months prior with 10⁶ PFU of VACV-WR (n = 3) or vGK5 (n = 4). (D) VACV titers were determined in organs 5 days post infection by the i.p. route and expressed as log₁₀ PFU per gram of lung and spleen tissue and PFU/ovary. – represents median values of titers in respective organs. N.S. = Not significant. P values were determined by Student's <i>t</i> test.</p

Cytokine responses and cytolytic activity in target organs.

<p>(A) IFN-γ responses of splenocytes from intranasally infected mice were measured in response to 3 VACV-specific CD8 T cell peptides (1 µg/ml) in an Elispot assay. Assays were performed using triplicate wells for each condition and individual mice/group. * = no responses detected. Seven days post infection, (B) splenocytes and (C) lung lymphocytes were isolated and CTL assays were carried out using RMA cells infected with VACV-WR (moi = 5), vGK5 (moi = 5) at different effector to target (E/T) ratios. Data shown are 1of 2 experiments performed. (D) PRNT50 antibody titers were measured in the sera of mice immunized 5 months prior with varying doses of vGK5 or 10^3.5 PFU of VACV-WR (n = 5/group).</p

Mice immunized with vGK5 are protected from a lethal challenge with VACV-WR.

<p>Mice were infected with 10^4.5, 10⁶ PFU of vGK5 (n = 3/group) by the intranasal route. 1 month later immunized mice and age matched naïve controls were challenged with a lethal dose of VACV-WR (10⁶ PFU) by the i.n. route. (A) Weights of mice were monitored over 5 days. Viral titers were measured in the (B) lungs and (C) ovaries and expressed as viral titers/gm lung tissue or ovaries. Each symbol represents the titer obtained in target organs of individual mice.</p