Differential Phosphorylation of Akt and signaling in CD4+ T Cells in Pathogenic and Apathogenic SIV Infection.

Increased CD4+ T cell apoptosis and activation induced cell death (AICD) as a result of HIV infection in humans and SIV infection in Rhesus macaques (RM) is indicative of disease. Some non-human primate species naturally infected by SIV, such as African sooty mangabeys (SM), do not succumb to SIV despite high viral loads. Previously, we showed that mRNA levels of GSK-3ß a kinase involved in T cell signaling, are significantly decreased in SIV+ RM compared to SIV+ SM. The current study confirms that expression of GSK-3ß is decreased at the protein level in SIV+ RM. In addition, CD4+ T cells from SIV+ RM, but not other animals show an increase in both total Akt, a kinase directly interacting with GSK-3ß and p-AktThr308 in response to stimulation via CD3/CD28, which is associated with an increase in apoptosis. Furthermore, the differences between the uninfected and pathogenically or non-pathogenically infected animals are not only species specific, but also T cell subset specific and that these trends correlate with AICD. This is one of few studies indicating the activity of Akt can be specific to only one phosphorylation site and may be linked to the differences in AICD and resistance to the lentivirus induced disease.

Prevalence of Mycoplasma hominis and Ureaplasma urealyticum in women undergoing an initial infertility evaluation.

AIMS: Mycoplasma hominis and Ureaplasma urealyticum are potentially pathogenic bacterial species that are frequently isolated from the urogenital tract of women. These pathogens could be responsible for various genitourinary diseases and have been associated with adverse pregnancy outcomes and female fertility problems. The aim of this study was to analyse the presence of M. hominis and U. urealyticum in the cervical canal of uterus of women with and without fertility problems. METHODS: Endocervical swabs obtained from women with reproductive problems and fertile women were tested by both cultivation and polymerase chain reaction. The antimicrobial susceptibility to the azithromycin, ciprofloxacin, doxycycline and erythromycine of the isolated strains of M. hominis and U. urealyticum was also tested by the microdilution broth method. RESULTS: A total of 111 women with fertile problems were examined. U. urealyticum was detected in samples from 44 (39.6%) women. M. hominis was detected in significantly fewer samples, i.e. only from 9 (8.1%) samples. From these, 6 (5.4%) women were positive for both microorganisms. The fertile group consisted from 23 women. The presence of U. urealyticum was detected in 8 (34.7%) of them. M. hominis was detected only in the mixture with U. urealyticum in 3 (13.0%) cases. The most effective antibiotic against both species in our study was doxycycline. CONCLUSION: The results show slightly higher incidence of M. hominis and U. urealyticum in the genitourinary tract of women with fertility problems compare with control group. The potential negative effect of these species on the reproduction ability of women was not observed.

[Infections caused by human alpha herpes viruses]. / Infekce vyvoláné lidskými alfa herpetickými viry.

Varicella-zoster virus (VZV), herpes simplex virus one (HSV-1) and herpes simplex virus two (HSV-2) represent three out of the eight known human herpesviruses and belong to the subfamily of α-herpesviruses. These viruses are present worldwide and humans are their sole host and reservoir. After the primary infection, these viruses persist in the body throughout life. The period of latency may be interrupted by reactivation of infection due to various factors. Each virus can induce a wide spectrum of diseases. The primary infection is typical for children and otherwise healthy individuals are often asymptomatic. It is mainly immunocompromised patients who are at risk of developing severe disease or complications when infected by these viruses. However, even in otherwise healthy individuals an infection by a-herpesviruses can run a severe course and lead to death.

[Opposite trends in the incidence of gonorrhea and syphilis in the East Bohemian Region - 30 years of surveillance]. / Protikladný vývoj trendu incidence kapavky a syfilis ve východoceském regionu - 30 let surveillance.

The epidemiology of selected sexually transmitted diseases in the Czech Republic has been carefully evaluated for many years. Data from 1981-2011 for eastern Bohemia shows a sharp decrease in the incidence of gonorrhea in 1993-1994 and a very low incidence thereafter with a slightly higher prevalence in males. However, syphilis and genitourinary infections with Chlamydia trachomatis show entirely opposite trends. Also, for the similar number of diagnostic tests performed, Chlamydia had a 10 fold higher rate of positive results. This underscores the changing epidemiology of sexually transmitted infections (STI) and necessity for adapting the reporting algorithms accordingly.

[An increase in the prevalence of syphilis in women in Eastern Bohemia - 30 years of surveillance]. / Rust prevalence syfilis u zen ve východoceském regionu - 30 let sledování

Epidemiological data on sexually transmitted infections in the Czech Republic has been carefully reported for many years. Here we present an analysis of regional data on syphilis spanning more than 30 years in eastern Bohemia. The epidemiological data were derived from the mandatory reporting of sexually transmitted diseases covering the period 1981-2011. The data showed a minimal incidence of cases in 1990 and an increasing trend thereafter. Two peaks in the incidence are apparent - within years 1995-1999 and 2003-2007. Interestingly, while before 1990 the numbers of positive men always exceeded those of women, their numbers equalized or even reversed within the two peaks. The results may also reflect trends in social change in the country after 1989. The analysis showed that the regional prevalence of syphilis in eastern Bohemia still remains low compared to some other regions of the Czech Republic.

[Legionella infection--a neglected problem]. / Legionelové infekce - opomíjený problém.

UNLABELLED: Pathogenic species of the Legionella genus can cause respiratory diseases ranging in severity from benign Pontiac fever to life-threatening Legionnaires disease often characterized by severe pneumonia, high fever, and multiple organ involvement. Predisposing underlying conditions, such as immunosuppression, chronic lung disease, and malignancies and other variables such as smoking and higher age constitute high-risk factors. Legionalla has been isolated from natural aquatic habitats (freshwater streams and lakes, water reservoirs, etc.), artificial sources, and also from humid soil. These pathogens are distributed worldwide. Besides water reservoirs (surface and underground water, fresh and salt water), they occur in a wide range of technical devices and systems - water distribution systems, showers, pools, spa systems, perlators, foggy makers, irrigation systems with sprinklers, cooling towers, etc. About 20% of detected Legionella infections in Europe have been associated with travel history. Travel-associated cases present a particular difficulty in terms of identifying the source of infection and implementing remedial measures. KEYWORDS: Legionella pneumophila - Legionnaires disease - Pontiac fever - nosocomial infections.

Ascorbic acid: an old player with a broad impact on body physiology including oxidative stress suppression and immunomodulation: a review.

Ascorbic acid is a low molecular weight antioxidant well known as anti-scorbut acting vitamin C in humans, primates and guinea pigs. This review summarizes basic data about ascorbic acid in its physiological action point of view. It is divided into biochemistry of ascorbic acid synthesis, mechanism of antioxidant action and participation in anabolism, pharmacokinetics and excretion, exogenous ascorbic acid immunomodulatory effect and participation in infectious diseases, impact on irradiation and intoxication pathogenesis, and supplementary demands. The primary intention was to consider ascorbic acid not only as an antioxidant but also as a chemical compound affecting multiple pathways with a potential beneficial impact in many diseases and processes in human body.

[Polio news]. / Novinky na poli poliomyelitidy--krátké sdelení.

Although the Word Health Organization (WHO) launched the global polio eradication initiative 22 years ago, reports of new cases of this serious disease from various parts of the world are far from being uncommon. Former Czechoslovakia became the first country in the world to achieve polio eradication at the nationwide level. Since summer 1960, no case of non-imported paralytic poliomyelitis has been reported since 1961, Czechoslovakia has been the first country in the world to control permanently the spread of wild poliovirus strains.

Macrophage-assisted inflammation and pharmacological regulation of the cholinergic anti-inflammatory pathway.

Macrophages play an important role in the immune system. They also participate in multiple processes including angiogenesis and triggering of inflammation. The present study summarizes pieces of knowledge on the importance of macrophages in disease, especially the inflammation. Special attention is paid to the cholinergic anti-inflammatory pathway (CAP) associated with the nicotinic acetylcholine receptor (nAChR) and the parasympathetic nervous system. The current pharmacological effectiveness in suppressing the inflammation in general and the septic shock in particular, is limited. CAP was discovered recently and it seems to be a suitable target for the development of new drugs. Moreover, available drugs binding to either nAChR or acetylcholinesterase (AChE) are candidates for either an inhibition or enhancement of CAP. Though the current scientific databases do not include all necessary data on the association of CAP with body functions and the research is quite intensive, the objective of the present review is to introduce the current trends and to critically evaluate CAP and macrophage-associated pathways.

Molecular epidemiology of varicella zoster virus.

Varicella zoster virus has highly conserved genome 125,000 base pairs. The different molecular genetic methods of analyzing VZV genome are discussed, as well as their results with regards to the virus phylogenesis, geographic distributions, possible recombination and virulence of different VZV strains.

[Herpes zoster in the Czech Republic--epidemiology and clinical manifestations]. / Herpes zoster v Ceské republice--epidemiologie a klinické projevy.

Herpes zoster (shingles) is a viral infection of the skin that manifests itself as painful, unilateral vesicular rash. The causative agent is varicella-zoster virus (VZV). Primary infection with VZV causes chickenpox, a common childhood infection, and then the virus lies dormant in the sensory neural ganglia, reactivating to cause shingles. The most important complications are neurological disorders (in particular postherpetic neuralgia) and eye disorders. First-line therapy are antiviral agents. A single vaccine has been registered to date. Herpes zoster occurs sporadically in the Czech Republic and its incidence is long-term stable. In 1990-2008 the average annual incidence was 6306 cases (61.3 cases/100,000 population), with the lowest number of 5511 cases (53.5/100,000) reported in 1991 and the highest number of 6,894 cases (67.6/100,000) reported in 2002. The incidence rate in females (69.9/100,000) was 1.4 times as high as in males (49.5/100,000). From the age perspective, the elderly are at a considerably higher risk of developing shingles. In 2008, the incidence rate was the highest in the age group 70 years (155.0/100,000). Nevertheless, the beginning of the upward trend is seen in the age group 45-49 years. Herpes zoster does not show any seasonal trend.

IL-15 is superior to IL-2 in the generation of long-lived antigen specific memory CD4 and CD8 T cells in rhesus macaques.

Using tetanus toxoid (TT) and influenza (Flu) immunization of rhesus macaques as a model, the effect of IL-2 and IL-15 on the generation and maintenance of antigen specific memory T cells was evaluated following primary and secondary immunization. Daily cytokine administration expanded primarily effector but not memory cells, while spacing cytokine administration to q3-7 days markedly enhanced TT and Flu specific memory responses. Following primary immunization, TT specific CD4 and influenza matrix protein (Flu-MP) specific CD8 effector responses were enhanced by IL-2 administration but CD8 specific memory responses were no different from cytokine non-treated monkeys. In contrast, expansion of Flu specific CD8 cells with IL-15 was only modest but resulted in significantly elevated levels of memory cells at 6 months. IL-15 also significantly enhanced early and late TT specific CD4 responses. The highest levels of primary effector and memory T cells were observed following alternate administration of both IL-2 and IL-15. Following booster immunization, however, only IL-15 appeared able to enhance CD8 T cell responses while IL-2 or IL2/IL-15 administration were less effective.

Norepinephrine enhances adhesion of HIV-1-infected leukocytes to cardiac microvascular endothelial cells.

Recent reports have indicated that norepinephrine (NE) enhances HIV replication in infected monocytes and promotes increased expression of select matrix metalloproteinases associated with dilated cardiomyopathy (DCM) in vitro in co-cultures of HIV-infected leukocytes and human cardiac microvascular endothelial cells (HMVEC-C). The influence of NE on HIV infection and leukocyte-endothelial interactions suggests a pathogenic role in AIDS-related cardiovascular disease. This study examined the effects of norepinephrine (NE) and HIV-1 infection on leukocyte adhesion to HMVEC-C. Both flow and static conditions were examined and the expression of selected adhesion molecules and cytokines were monitored in parallel. NE pretreatment resulted in a detectable, dose-dependent increase of leukocyte-endothelial adhesion (LEA) with both HIV-1-infected and -uninfected peripheral blood mononuclear cells (PBMCs) relative to media controls after 48 hr in co-culture with HMVEC-C in vitro. However, the combination of NE plus HIV infection resulted in a significant (P < 0.0001) 18-fold increase in LEA over uninfected media controls. Increased levels in both cell-associated and -soluble ICAM-1 and E-Selectin but not VCAM-1 correlated with increased LEA and with HIV-1 infection or NE pretreatment. Blocking antibodies specific for ICAM-1 or E-Selectin inhibited HIV-NE-induced LEA. These data suggest a model in which NE primes HIV-1-infected leukocytes for enhanced adhesion and localization in HMVEC-C where they can initiate and participate in vascular injury associated with AIDS-related cardiomyopathy.

Administration of recombinant rhesus interleukin-12 during acute simian immunodeficiency virus (SIV) infection leads to decreased viral loads associated with prolonged survival in SIVmac251-infected rhesus macaques.

The ability of recombinant rhesus interleukin-12 (rMamu-IL-12) administration during acute simian immunodeficiency virus SIVmac251 infection to influence the quality of the antiviral immune responses was assessed in rhesus macaques. Group I (n = 4) was the virus-only control group. Group II and III received a conditioning regimen of rMamu-IL-12 (10 and 20 microg/kg, respectively, subcutaneously [s.c.]) on days -2 and 0. Thereafter, group II received 2 microg of IL-12 per kg and group III received 10 microg/kg s.c. twice a week for 8 weeks. On day 0 all animals were infected with SIVmac251 intravenously. While all four group I animals and three of four group II animals died by 8 and 10 months post infection (p.i.), all four group III animals remained alive for >20 months p.i. The higher IL-12 dose led to lower plasma viral loads and markedly lower peripheral blood mononuclear cell and lymph node proviral DNA loads. During the acute viremia phase, the high-IL-12-dose monkeys showed an increase in CD3(-) CD8 alpha/alpha(+) and CD3(+) CD8 alpha/alpha(+) cells and, unlike the control and low-IL-12-dose animals, did not demonstrate an increase in CD4(+) CD45RA(+) CD62L(+) naive cells. The high-IL-12-dose animals also demonstrated that both CD8 alpha/alpha(+) and CD8 alpha/beta(+) cells produced antiviral factors early p.i., whereas only CD8 alpha/beta(+) cells retained this function late p.i. Long-term survival correlated with sustained high levels of SIV gag/pol and SIV env cytotoxic T lymphocytes and retention of high memory responses against nominal antigens. This is the first study to demonstrate the capacity of IL-12 to significantly protect macaques from SIV-induced disease, and it provides a useful model to more precisely identify correlates of virus-specific disease-protective responses.

Identification of protein kinases dysregulated in CD4(+) T cells in pathogenic versus apathogenic simian immunodeficiency virus infection.

Human immunodeficiency virus infection in humans and simian immunodeficiency virus (SIV) infection in rhesus macaques (RM) leads to a generalized loss of immune responses involving perturbations in T-cell receptor (TCR) signaling. In contrast, naturally SIV-infected sooty mangabeys (SM) remain asymptomatic and retain immune responses despite relatively high viral loads. However, SIV infection in both RM and SM led to similar decreases in TCR-induced Lck phosphorylation. In this study, a protein tyrosine kinase (PTK) differential display method was utilized to characterize the effects of in vivo SIV infection on key signaling molecules of the CD4(+) T-cell signaling pathways. The CD4(+) T cells from SIV-infected RM, but not SIV-infected SM, showed chronic downregulation of baseline expression of MLK3, PRK, and GSK3, and symptomatically SIV-infected RM showed similar downregulation of MKK3. In vitro TCR stimulation with or without CD28 costimulation of CD4(+) T cells did not lead to the enhancement of gene transcription of these PTKs. While the CD4(+) T cells from SIV-infected RM showed a significant increase of the baseline and anti-TCR-mediated ROR2 transcription, SIV infection in SM led to substantially decreased anti-TCR-stimulated ROR2 transcription. TCR stimulation of CD4(+) T cells from SIV-infected RM (but not SIV-infected SM) led to the repression of CaMKKbeta and the induction of gene transcription of MLK2. Studies of the function of these molecules in T-cell signaling may lead to the identification of potential targets for specific intervention, leading to the restoration of T-cell responses.

Cloning, sequencing, and homology analysis of nonhuman primate Fas/Fas-ligand and co-stimulatory molecules.

The finding that a single administration of select recombinant human cytokines to nonhuman primates leads to potent cytokine-neutralizing antibody responses in the heterologous host despite >95% homology at the nucleotide and protein level prompted our laboratory to clone, sequence, and prepare recombinant nonhuman primate cytokines, chemokines, growth factors, and other immunoregulatory molecules. In the present report, we present findings on the gene sequences encoding the nonhuman primate homologues of human CD80, CD86, their ligands CD28 and CD152, CD154, CD95, and CD95-L from rhesus macaques and for phylogenetic analysis from pig-tailed macaques, African sooty mangabey monkeys, baboons, and vervets as well as select molecules from the New World aotus and marmoset monkeys. With the exception of CD95, the homology between nonhuman primate and human co-stimulatory molecules was above 95%. In contrast, CD95 was only 89.2% homologous to human CD95, but the differences were essentially found in the transmembrane and intracellular (death) domains. The extracellular portion of CD95 was more homologous which was in accordance with approximately 98% homology between Old World monkey and human CD95-L. In general, sequences from the New World monkey species appeared equidistant to sequences from Old World species and humans in terms of homology suggesting distinct evolutionary patterns. Of interest was the isolation of various splice variants of monkey CD86, CD152 (CTLA-4), CD154, and CD95 transcripts. This is also the first report documenting the occurrence of natural CD86 variants with deleted transmembrane domains, found both in sooty mangabeys and baboon RNA samples. Monkey CD95 showed various deletions and addition of residues in the transmembrane and intracytoplasmic domains compared with human CD95 and between Old and New World species. Subcloning of rhesus CD154 into an expression vector demonstrated expression of a functional protein in cell culture. The other genes are being cloned into expression vectors for the preparation and biological characterization of the nonhuman primate molecules. These investigations will provide novel reagents for in vivo use as immunomodulatory reagents in nonhuman primates in studies which may provide a rationale for their use in humans.

Failure to expand influenza and tetanus toxoid memory T cells in vitro correlates with disease course in SIV infected rhesus macaques.

Marked decreases in influenza (flu) and tetanus toxoid (T.T.) antigen specific CD8(+) and CD4(+) T cell memory responses were noted shortly after SIV infection in monkeys that go on to develop clinical disease within 18 months (normal progressor, NP) following SIV infection but not in monkeys that remain asymptomatic >3 years post SIV infection (long-term nonprogressor, LTNP). While PBMCs from NP and LTNP monkeys demonstrate both low and high avidity flu and T.T. specific CD8(+) and CD4(+)T cell immune responses prior to SIV infection, the PBMCs from NP but not LTNP fail to generate high avidity T cell responses post SIV infection. This failure to generate high avidity T cell responses in vitro correlated with increased apoptotic cell death in PBMC cultures from NP animals. Since high avidity antigen specific CTLs have been shown to be most efficient in eliminating viral infections, the present finding has important implications for the evaluation of the level of immune reconstitution following various modalities of therapy in HIV-1 infected patients.

Effects of norepinephrine, HIV type 1 infection, and leukocyte interactions with endothelial cells on the expression of matrix metalloproteinases.

The expression of matrix metalloproteinases (MMPs) associated with AIDS-related cardiomypathies and cocaine abuse was examined in an in vitro coculture model. Human peripheral blood mononuclear cells (PBMCs), HIV infected or uninfected, were placed in coculture with primary human cardiac microvascular endothelial cells (HMVEC-C) in the presence or absence of the cocaine-inducible catecholamine norepinephrine (NE). Culture supernatants were assayed for MMP-1, -2, -3, -7, -9, and -13, and for tissue inhibitor of metalloproteinase 1 (TIMP-1) and TIMP-2, by enzyme-linked immunosorbent assay. Low levels of constitutively expressed MMP-1 and -2 were detected in individual cultures of HMVEC-C and PBMCs. NE did not induce MMP or TIMP expression by HMVEC-C and caused modest increases (3- to 4-fold) in MMP-1 and -2 by uninfected PBMCs. Increased levels of NE-induced MMP-1 (5-fold) and MMP -2 (15-fold) were detected in cocultures of HMVEC-C and uninfected PBMCs. HIV infection enhanced MMP-1 (46-fold) and MMP-2 (48-fold) and active MMP-7 (33-fold) and MMP-9 (50-fold) by PBMCs. Coculture of HIV-infected PBMCs with HMVEC-C increased MMP-1 (110-fold) and MMP-2 (307-fold) but not active MMP-7 and -9. The combination of NE, HIV infection, and coculture increased MMP-1 (126-fold) and MMP-2 (467-fold), and active MMP-7 (65-fold) and MMP-9 (75-fold). MMP-3 or-13 was not detected in any of the treatment groups and TIMP-1 and -2 appeared inversely proportional to the observed levels of MMPs. These results suggest that HIV infection, NE, and leukocyte endothelial interactions demonstrate separate and overlapping cooperative effects on the regulation of expression of TIMPs and MMPs associated with AIDS-related cardiomyopathies.

Relative resistance in the development of T cell anergy in CD4+ T cells from simian immunodeficiency virus disease-resistant sooty mangabeys.

Despite high viral loads, T cells from sooty mangabey (SM) monkeys that are naturally infected with SIV but remain clinically asymptomatic, proliferate and demonstrate normal Ag-specific memory recall CD4(+) T cell responses. In contrast, CD4(+) T cells from rhesus macaques (RM) experimentally infected with SIV lose Ag-specific memory recall responses and develop immunological anergy. To elucidate the mechanisms for these distinct outcomes of lentiviral infection, highly enriched alloreactive CD4(+) T cells from humans, RM, and SM were anergized by TCR-only stimulation (signal 1 alone) and subsequently challenged with anti-CD3/anti-CD28 Abs (signals 1 + 2). Whereas alloreactive CD4(+)T cells from humans and RM became anergized, surprisingly, CD4(+) T cells from SM showed marked proliferation and IL-2 synthesis after restimulation. This resistance to undergo anergy was not secondary to a global deficiency in anergy induction of CD4(+) T cells from SM since incubation of CD4(+) T cells with anti-CD3 alone in the presence of rapamycin readily induced anergy in these cells. The resistance to undergo anergy was reasoned to be due to the ability of CD4(+) T cells from SM to synthesize IL-2 when incubated with anti-CD3 alone. Analysis of phosphorylated kinases involved in T cell activation showed that the activation of CD4(+) T cells by signal 1 in SM elicited a pattern of response that required both signals 1 + 2 in humans and RM. This function of CD4(+) T cells from SM may contribute to the resistance of this species to SIV-induced disease.

Induction of long-term protective effects against heterologous challenge in SIVhu-infected macaques.

A group of three rhesus macaques were inoculated with SIV isolated from a human (SIVhu) accidentally exposed and infected with SIVsm. Extensive sequence analyses of SIVhu obtained from the human and macaques following infection indicated the presence of truncated nef. Not only did nef fail to repair itself in vivo postinfection (p.i.), but instead, further mutations added additional stop codons with increasing time p.i. Infection of these animals was associated with minimal acute viral replication, followed by undetectable plasma viral loads and only intermittent PCR detection up to 5 years p.i. The three SIVhu infected and three control monkeys were then challenged with the heterologous highly pathogenic SHIV89.6p. All three controls became infected and showed rapid declines in peripheral CD4(+) lymphocytes, disease, and death at 10 and 32 weeks p.i., respectively. In contrast, all three animals previously infected with SIVhu are healthy and exhibit stable CD4(+) lymphocyte levels and undetectable plasma viral loads at >20 months post-SHIV89. 6p challenge. Only transient, low levels of SHIV replication were noted in these animals. Whereas responses to SIVgag/pol were noted, no evidence for SIV/SHIV envelope cross-reactivity was detected by antibody or CTL analyses, suggesting that the protective immune mechanisms to the heterologous challenge isolate were most likely not directed to envelope but rather to other viral determinants.