Captopril mitigates splenomegaly and myelofibrosis in the Gata1low murine model of myelofibrosis.

Allogeneic stem cell transplantation is currently the only curative therapy for primary myelofibrosis (MF), while the JAK2 inhibitor, ruxolitinib. Has been approved only for palliation. Other therapies are desperately needed to reverse life-threatening MF. However, the cell(s) and cytokine(s) that promote MF remain unclear. Several reports have demonstrated that captopril, an inhibitor of angiotensin-converting enzyme that blocks the production of angiotensin II (Ang II), mitigates fibrosis in heart, lung, skin and kidney. Here, we show that captopril can mitigate the development of MF in the Gata1low mouse model of primary MF. Gata1low mice were treated with 79 mg/kg/d captopril in the drinking water from 10 to 12 months of age. At 13 months of age, bone marrows were examined for fibrosis, megakaryocytosis and collagen expression; spleens were examined for megakaryocytosis, splenomegaly and collagen expression. Treatment of Gata1low mice with captopril in the drinking water was associated with normalization of the bone marrow cellularity; reduced reticulin fibres, splenomegaly and megakaryocytosis; and decreased collagen expression. Our findings suggest that treating with the ACE inhibitors captopril has a significant benefit in overcoming pathological changes associated with MF.

Suppression of transforming growth factor ß receptor 2 and Smad5 is associated with high levels of microRNA miR-155 in the oral mucosa during chronic simian immunodeficiency virus infection.

Chronic human immunodeficiency virus and simian immunodeficiency virus (HIV and SIV) infections are characterized by mucosal inflammation in the presence of anti-inflammatory cytokines such as transforming growth factor ß (TGFß). The mechanisms for refractiveness to TGFß are not clear. Here we show that the expression of microRNA miR-155 was significantly upregulated in the oropharyngeal mucosa during chronic SIV infection and was coincident with downregulation of TGFß receptor 2 (TGFß-R2) and SMAD5, key TGFß signaling genes that harbor putative target sites for miR-155. Ectopic expression of miR-155 in vitro was found to significantly downregulate TGFß-R2 and Smad5 expression, suggesting a role for miR-155 in the suppression of TGFß-R2 and SMAD5 genes in vivo. The downregulation of TGFß signaling genes by miR-155 likely contributes to the nonresponsiveness to TGFß during SIV infection and may inadvertently aid in increased immune activation during HIV and SIV infections.

Early treatment with reverse transcriptase inhibitors significantly suppresses peak plasma IFNα in vivo during acute simian immunodeficiency virus infection.

Innate interferons (IFN) are comprised of multiple Type I and III subtypes. The in vivo kinetics of subtype responses during human immunodeficiency virus (HIV) infection is not well defined. Using the acute simian immunodeficiency virus (SIV) infection model, we show that plasma IFNα levels peak at day 10 post-infection (pi) after which they rapidly declined. The mRNA expression of Type I and III IFN subtypes were significantly elevated in the lymph nodes (LN) at day 10 pi. Though the expression levels of all subtypes declined by day 14-31 pi, numerous subtypes remained elevated suggesting that ongoing viral replication in LN continues to drive induction of these subtypes. Interestingly, treatment with reverse transcriptase (RT) inhibitors at day 7 pi significantly suppressed plasma IFNα responses by day 10 pi that significantly correlated with cell-associated SIV DNA loads suggesting that RT byproducts such as viral DNA likely plays a role in driving IFN responses during acute SIV infection. Quantification of Type I and III subtype transcripts in sorted subsets of LN CD4+ and CD8+ T cells, CD14+/CD14- monocytes/macrophages, and total CD11c/CD123+ dendritic cells (DC) at day 10 pi showed that DC expressed ∼3-4 log more subtype transcripts as compared to the other subsets. Taken together, our results provide new insights into the kinetics of innate interferon responses during early stages of infection, and provide evidence that DC's are a major in vivo source of innate IFN during acute SIV infection.

Captopril reduces lung inflammation and accelerated senescence in response to thoracic radiation in mice.

The lung is sensitive to radiation and exhibits several phases of injury, with an initial phase of radiation-induced pneumonitis followed by delayed and irreversible fibrosis. The angiotensin-converting enzyme inhibitor captopril has been demonstrated to mitigate radiation lung injury and to improve survival in animal models of thoracic irradiation, but the mechanism remains poorly understood. Here we investigated the effect of captopril on early inflammatory events in the lung in female CBA/J mice exposed to thoracic X-ray irradiation of 17-17.9 Gy (0.5-0.745 Gy min-1). For whole-body + thoracic irradiation, mice were exposed to 7.5 Gy (0.6 Gy min-1) total-body 60Co irradiation and 9.5 Gy thoracic irradiation. Captopril was administered orally (110 mg kg-1 day-1) in the drinking water, initiated 4 h through to150 days post-irradiation. Captopril treatment increased survival from thoracic irradiation to 75% at 150 days compared with 0% survival in vehicle-treated animals. Survival was characterized by a significant decrease in radiation-induced pneumonitis and fibrosis. Investigation of early inflammatory events showed that captopril significantly attenuated macrophage accumulation and decreased the synthesis of radiation-induced interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α) pro-inflammatory cytokines in the lungs of irradiated mice. Suppression of IL-1ß and TNF-α correlated with an increase of the anti-inflammatory cytokine IL-10 in the spleen with captopril treatment. We also found that captopril decreased markers for radiation-induced accelerated senescence in the lung tissue. Our data suggest that suppression of inflammation and senescence markers, combined with an increase of anti-inflammatory factors, are a part of the mechanism for captopril-induced survival in thoracic irradiated mice.

Elucidation of Antimicrobial Susceptibility Profiles and Genotyping of Salmonella enterica Isolates from Clinical Cases of Salmonellosis in New Mexico in 2008.

In this study, we investigated the antimicrobial susceptibility profiles and the distribution of some well known genetic determinants of virulence in clinical isolates of Salmonella enterica from New Mexico. The minimum inhibitory concentrations (MICs) for various antimicrobials were determined by using the E-test strip method according to CLSI guidelines. Virulence genotyping was performed by polymerase chain reaction (PCR) using primers specific for known virulence genes of Salmonella enterica. Of 15 isolates belonging to 11 different serovars analyzed, one isolate of Salmonella Typhimurium was resistant to multiple drugs namely ampicillin, amoxicillin / clavulanic acid, chloramphenicol and tetracycline, that also harbored class 1 intergron, bla(TEM) encoding genes for ß-lactamase, chloramphenicol acetyl transferase (cat1), plus floR, tet(C) and tet(G). This strain was phage typed as DT104. PCR analysis revealed the presence of invA, hilA, stn, agfA and spvR virulence genes in all the isolates tested. The plasmid-borne pefA gene was absent in 11 isolates, while 5 isolates lacked sopE. One isolate belonging to serogroup E4 (Salmonella Sombre) was devoid of multiple virulence genes pefA, iroB, shdA and sopE. These results demonstrate that clinical Salmonella serotypes from New Mexico used here are predominantly sensitive to multiple antimicrobial agents, but vary in their virulence genotypes. Information on antimicrobial sensitivity and virulence genotypes will help in understanding the evolution and spread of epidemic strains of Salmonella enterica in the region of study.

Prevalence and heterogeneity of hemolysin gene vhh among hatchery isolates of Vibrio harveyi in India.

Vibrio harveyi, pathogenic to fish, harbor a hemolysin gene vhh, the homologues of which are found in many species of the Genus Vibrio. In this study, we investigated the prevalence of vhh gene among V. harveyi isolated from Penaeus monodon hatcheries in India by polymerase chain reaction (PCR). The vhh was detected in 67 of the 70 V. harveyi isolates tested in this study using different combinations of PCR primers. A variant vhh gene detected in a minority of strains was cloned, sequenced, and the recombinant protein was expressed in Escherichia coli. The deduced amino acid sequence of the cloned gene was 86% similar to the previously reported amino acid sequences of VHH. The results of this study suggest that though V. harveyi strains invariably harbor vhh, the sequence variants of the hemolysin gene exist that may impede their detection by PCR.

Gender differences in innate responses and gene expression profiles in memory CD4 T cells are apparent very early during acute simian immunodeficiency virus infection.

Gender differences in Human immunodeficiency virus (HIV) disease progression and comorbidities have been extensively reported. Using the simian immunodeficiency virus (SIV) infected rhesus macaque model, we show that these differences are apparent very early during the course of infection. Though there were no major changes in the proportions of CD4 T cells or its subsets, central memory CD4 T cells from female macaques were found to differentially regulate a significantly larger number of genes at day 4 post-infection (PI) as compared to males. Pathway analysis revealed divergence of both canonical and biological pathways that persisted at day 10 PI. Changes in gene expression profiles were accompanied by a significant increase in plasma levels of pro-inflammatory mediators such as MCP-1/CCL2, I-TAC/CXCL11, and MIF. Though plasma levels of IFNα did not differ between male and female macaques, the expression levels of IFNα subtype-14, 16, IFNß, and IFNω were significantly upregulated in the lymph nodes of female macaques at day 10 PI as compared to male macaques. Our results suggest that the pathogenic sequelae seen during chronic infection may be shaped by gender differences in immune responses induced very early during the course of HIV infection.

Interferon-α Subtypes As an Adjunct Therapeutic Approach for Human Immunodeficiency Virus Functional Cure.

Human immunodeficiency virus (HIV) establishes life-long latency in infected individuals. Although highly active antiretroviral therapy (HAART) has had a significant impact on the course of HIV infection leading to a better long-term outcome, the pool of latent reservoir remains substantial even under HAART. Numerous approaches have been under development with the goal of eradicating the latent HIV reservoir though with limited success. Approaches that combine immune-mediated control of HIV to activate both the innate and the adaptive immune system under suppressive therapy along with "shock and kill" drugs may lead to a better control of the reactivated virus. Interferon-α (IFN-α) is an innate cytokine that has been shown to activate intracellular defenses capable of restricting and controlling HIV. IFN-α, however, harbors numerous functional subtypes that have been reported to display different binding affinities and potency. Recent studies have suggested that certain subtypes such as IFN-α8 and IFN-α14 have potent anti-HIV activity with little or no immune activation, whereas other subtypes such as IFN-α4, IFN-α5, and IFN-α14 activate NK cells. Could these subtypes be used in combination with other strategies to reduce the latent viral reservoir? Here, we review the role of IFN-α subtypes in HIV infection and discuss the possibility that certain subtypes could be potential adjuncts to a "shock and kill" or therapeutic vaccination strategy leading to better control of the latent reservoir and subsequent functional cure.

Prior Exposure to Zika Virus Significantly Enhances Peak Dengue-2 Viremia in Rhesus Macaques.

Structural and functional homologies between the Zika and Dengue viruses' envelope proteins raise the possibility that cross-reactive antibodies induced following Zika virus infection might enhance subsequent Dengue infection. Using the rhesus macaque model we show that prior infection with Zika virus leads to a significant enhancement of Dengue-2 viremia that is accompanied by neutropenia, lympocytosis, hyperglycemia, and higher reticulocyte counts, along with the activation of pro-inflammatory monocyte subsets and release of inflammatory mediators. Zika virus infection induced detectable Dengue cross-reactive serum IgG responses that significantly amplified after Dengue-2 virus infection. Serum from Zika virus immune animals collected prior to Dengue-2 infection showed significant capacity for in vitro antibody dependent enhancement of Dengue-1, 2, 3 and 4 serotypes suggesting that pre-existing immunity to Zika virus could potentially enhance infection by heterologous Dengue serotypes. Our results provide first in vivo evidence that prior exposure to Zika virus infection can enhance Dengue infection, which has implications for understanding pathogenesis and the development of vaccines.

Significant Depletion of CD4(+) T Cells Occurs in the Oral Mucosa during Simian Immunodeficiency Virus Infection with the Infected CD4(+) T Cell Reservoir Continuing to Persist in the Oral Mucosa during Antiretroviral Therapy.

Human and simian immunodeficiency virus (HIV and SIV) infections are characterized by manifestation of numerous opportunistic infections and inflammatory conditions in the oral mucosa. The loss of CD4(+) T cells that play a critical role in maintaining mucosal immunity likely contributes to this process. Here we show that CD4(+) T cells constitute a minor population of T cells in the oral mucosa and display a predominantly central memory phenotype mirroring other mucosal sites such as the rectal mucosa. Chronic SIV infection was associated with a near total depletion of CD4(+) T cells in the oral mucosa that appear to repopulate during antiretroviral therapy (ART). Repopulating CD4(+) T cells harbored a large fraction of Th17 cells suggesting that ART potentially reconstitutes oral mucosal immunity. However, a minor fraction of repopulating CD4(+) T cells harbored SIV DNA suggesting that the viral reservoir continues to persist in the oral mucosa during ART. Therapeutic approaches aimed at obtaining sustainable CD4(+) T cell repopulation in combination with strategies that can eradicate the latent viral reservoir in the oral mucosa are essential for better oral health and long-term outcome in HIV infected patients.

Rhesus macaque lymph node PD-1(hi)CD4+ T cells express high levels of CXCR5 and IL-21 and display a CCR7(lo)ICOS+Bcl6+ T-follicular helper (Tfh) cell phenotype.

CD4 T follicular helper (Tfh) cells play a unique and essential role in the generation of B cell responses in the lymph node microenvironment. Here we sought to determine if differential expression of PD-1 could be used to delineate Tfh cells in rhesus macaque lymph nodes (LN). CD3(+)CD4(+) T cells were found to harbor a unique subset of cells that expressed the Program death-1 (PD-1) receptor at significantly high levels that were enriched in the LN compartment as compared to peripheral blood. The LN CD4(+)PD1(hi) T cells expressed a predominantly CD28(+)CD95(+) central memory phenotype and were CCR7(lo)ICOS(hi)Bcl6(hi). Additionally, CD4(+)PD1(hi) T cells preferentially expressed high levels of CXCR5 and IL-21 and significantly correlated with Bcl6(+)Ki-67(+) IgG(+) B cells. As Bcl6 is primarily expressed by proliferating B cells within active germinal centers, our results suggest that LN CD4(+)PD1(hi) T cells likely localize to active GC regions, a characteristic that is attributable to Tfh cells. Overall, our findings suggest that high levels of PD-1 expression on CD4(+) T cells in LN of rhesus macaques can serve as a valuable marker to identify Tfh cells and has implications for studying the role of Tfh cells in Human immunodeficiency virus (HIV), Simian immunodeficiency virus (SIV) and other infectious diseases that use the rhesus macaque model.

Early short-term antiretroviral therapy is associated with a reduced prevalence of CD8(+)FoxP3(+) T cells in simian immunodeficiency virus-infected controller rhesus macaques.

Regulatory T cells contain a mix of CD4 and CD8 T cell subsets that can suppress immune activation and at the same time suppress immune responses, thereby contributing to disease progression. Recent studies have shown that an increased prevalence of CD8(+)FoxP3(+) T regulatory cells was associated with immune suppression and diminished viral control in simian immunodeficiency virus (SIV)-infected rhesus macaques. Preventing an increase in the prevalence of CD8 T regulatory subsets is likely to lead to a better long-term outcome. Here we show that short-term antiretroviral therapy initiated within 1 week after SIV infection was associated with lower viral set point and immune activation after withdrawal of therapy as compared to untreated animals. Early short-term treated controller animals were found to have better SIV-specific immune responses and a significantly lower prevalence of immunosuppressive CD8(+)FoxP3(+) T cells. Lower levels of CD8(+)FoxP3(+) T cells coincided with preservation of CD4(+)FoxP3(+) T cells at homeostatic levels, and significantly correlated with lower immune activation, suggesting a role for viral infection-driven immune activation in the expansion of CD8(+)FoxP3(+) T cells. Interestingly, initiation of continuous therapy later in infection did not reduce the increased prevalence of CD8(+)FoxP3(+) T cells to homeostatic levels. Taken together, our results suggest that early antiretroviral therapy preserves the integrity of the immune system leading to a lower viral set point in controller animals, and prevents alterations in the homeostatic balance between CD4(+) and CD8(+) T regulatory cells that could aid in better long-term outcome.