Glutamine supplementation suppresses herpes simplex virus reactivation.

Chronic viral infections are difficult to treat, and new approaches are needed, particularly those aimed at reducing reactivation by enhancing immune responses. Herpes simplex virus (HSV) establishes latency and reactivates frequently, and breakthrough reactivation can occur despite suppressive antiviral therapy. Virus-specific T cells are important to control HSV, and proliferation of activated T cells requires increased metabolism of glutamine. Here, we found that supplementation with oral glutamine reduced virus reactivation in latently HSV-1-infected mice and HSV-2-infected guinea pigs. Transcriptome analysis of trigeminal ganglia from latently HSV-1-infected, glutamine-treated WT mice showed upregulation of several IFN-γ-inducible genes. In contrast to WT mice, supplemental glutamine was ineffective in reducing the rate of HSV-1 reactivation in latently HSV-1-infected IFN-γ-KO mice. Mice treated with glutamine also had higher numbers of HSV-specific IFN-γ-producing CD8 T cells in latently infected ganglia. Thus, glutamine may enhance the IFN-γ-associated immune response and reduce the rate of reactivation of latent virus infection.

HLA-DQ ß1 alleles associated with Epstein-Barr virus (EBV) infectivity and EBV gp42 binding to cells.

Epstein-Barr virus (EBV) infects B cells and ~95% of adults are infected. EBV glycoprotein gp42 is essential for entry of virus into B cells. EBV gp42 binds to the ß1 chain of HLA-DQ, -DR, and -DP on B cells, and uses these molecules for infection. To investigate if certain HLA-DQ alleles are associated with EBV seronegativity, we recruited ~3,300 healthy adult blood donors, identified 106 EBV-seronegative individuals, and randomly selected a control group of EBV-seropositive donors from the donor pool. A larger than expected proportion of EBV-seronegative subjects were HLA-DQ ß1 *04/*05 and *06/*06, and to a lesser extent, *02/*03, compared with the control group, while a larger than expected portion of EBV-seropositive persons were HLA-DQ ß1 *02/*02. We examined the ability of EBV gp42 to bind to different HLA-DQ molecules using human and mouse cells stably expressing these alleles. EBV gp42 bound less effectively to cells expressing HLA-DQ ß1 *04/*05, *06/*06, or *03/*03 than to cells expressing HLA-DQ ß1 *02/*02. These data are consistent with our observations of increased EBV seronegativity with DQ ß1 *04/*05 or *06/*06 alleles. These findings emphasize the importance of a single genetic locus (HLA-DQ ß1) to influence infectivity with EBV.

Soluble rhesus lymphocryptovirus gp350 protects against infection and reduces viral loads in animals that become infected with virus after challenge.

Epstein-Barr virus (EBV) is a human lymphocryptovirus that is associated with several malignancies. Elevated EBV DNA in the blood is observed in transplant recipients prior to, and at the time of post-transplant lymphoproliferative disease; thus, a vaccine that either prevents EBV infection or lowers the viral load might reduce certain EBV malignancies. Two major approaches have been suggested for an EBV vaccine- immunization with either EBV glycoprotein 350 (gp350) or EBV latency proteins (e.g. EBV nuclear antigens [EBNAs]). No comparative trials, however, have been performed. Rhesus lymphocryptovirus (LCV) encodes a homolog for each gene in EBV and infection of monkeys reproduces the clinical, immunologic, and virologic features of both acute and latent EBV infection. We vaccinated rhesus monkeys at 0, 4 and 12 weeks with (a) soluble rhesus LCV gp350, (b) virus-like replicon particles (VRPs) expressing rhesus LCV gp350, (c) VRPs expressing rhesus LCV gp350, EBNA-3A, and EBNA-3B, or (d) PBS. Animals vaccinated with soluble gp350 produced higher levels of antibody to the glycoprotein than those vaccinated with VRPs expressing gp350. Animals vaccinated with VRPs expressing EBNA-3A and EBNA-3B developed LCV-specific CD4 and CD8 T cell immunity to these proteins, while VRPs expressing gp350 did not induce detectable T cell immunity to gp350. After challenge with rhesus LCV, animals vaccinated with soluble rhesus LCV gp350 had the best level of protection against infection based on seroconversion, viral DNA, and viral RNA in the blood after challenge. Surprisingly, animals vaccinated with gp350 that became infected had the lowest LCV DNA loads in the blood at 23 months after challenge. These studies indicate that gp350 is critical for both protection against infection with rhesus LCV and for reducing the viral load in animals that become infected after challenge. Our results suggest that additional trials with soluble EBV gp350 alone, or in combination with other EBV proteins, should be considered to reduce EBV infection or virus-associated malignancies in humans.

Evaluation of the safety and efficacy of micafungin in Japanese patients with deep mycosis: a post-marketing survey report.

The safety and efficacy of micafungin were evaluated in a Japanese post-marketing survey involving 1,142 patients with deep mycosis caused by Candida or Aspergillus. The overall clinical response was 83.0%, and the respective responses for patients with candidiasis or aspergillosis were 86.3 and 70.8%. With regard to drug reactions, 562 adverse reactions were observed in 28.5% of patients. Among the 83 serious adverse drug reactions reported by 53 patients, a causal relationship with micafungin was assessed as definite or probable for 6 reactions in 5 patients. Age and baseline hepatic and renal function status did not affect the incidence of adverse reactions, although incidence increased significantly in proportion to the severity of mycosis and daily dose (p < 0.01). In multiple logistic regression analysis, neither baseline hepatic impairment nor increased daily dose of micafungin affected the incidence of hepatobiliary disorders, however, the severity of mycosis was found to correlate significantly with hepatobiliary disorders (p = 0.031). Taken together, our post-marketing findings show that micafungin is effective against deep mycosis caused by Candida or Aspergillus in patients across a range of backgrounds.

Kinetics of Epstein-Barr virus load and virus-specific CD8+ T cells in acute infectious mononucleosis.

BACKGROUND: During the convalescent phase of acute infectious mononucleosis (AIM), Epstein-Barr virus (EBV) load shrinks rapidly in association with a rapid decline in the number of EBV-specific CD8(+) T cells. The actual contribution of EBV-specific CD8(+) T cells in reducing EBV load, however, is not known. OBJECTIVES: To clarify the impact of EBV-specific CD8(+) T cells on the contraction of EBV load in AIM, we estimated half-lives of both EBV load and EBV-specific CD8(+) T cells. STUDY DESIGN: Blood was serially taken from five pediatric patients with AIM during the convalescent period, including the very early phase, and both EBV load and EBV-specific CD8(+) T cell numbers were assayed. RESULTS: EBV load declined rapidly (half-life 1.5 d) during the first 2 weeks after onset of symptoms. This half-life was seven-fold shorter than that reported for CD27(+) memory B cells. Subsequently, the EBV load declined much more slowly, with a half-life of 38.7 d. EBV-specific CD8(+) T cell numbers also declined concomitantly with the decrease in EBV load. The half-life of EBV-specific CD8(+) T cells during first 2 weeks was 2.9 d. The number of EBV-specific CD8(+) T cells and the rate of change of viral load correlated significantly (R(2) ≥ 0.8; p ≤ 0.02). CONCLUSIONS: The short half-life of EBV load, together with the strong correlation between the number of EBV-specific CD8(+) T cells and the rate of change of viral load indicates an active role for EBV-specific CD8(+) T cells in elimination of EBV in AIM.

Long-term administration of valacyclovir reduces the number of Epstein-Barr virus (EBV)-infected B cells but not the number of EBV DNA copies per B cell in healthy volunteers.

Epstein-Barr virus (EBV) establishes a latent infection in B cells in the blood, and the latent EBV load in healthy individuals is generally stable over time, maintaining a "set point." It is unknown if the EBV load changes after long-term antiviral therapy in healthy individuals. We treated volunteers with either valacyclovir (valaciclovir) or no antiviral therapy for 1 year and measured the amount of EBV DNA in B cells every 3 months with a novel, highly sensitive assay. The number of EBV-infected B cells decreased in subjects receiving valacyclovir (half-life of 11 months; P = 0.02) but not in controls (half-life of 31 years; P = 0.86). The difference in the slopes of the lines for the number of EBV-infected B cells over time for the valacyclovir group versus the control group approached significance (P = 0.054). In contrast, the number of EBV DNA copies per B cell remained unchanged in both groups (P = 0.62 and P = 0.92 for the control and valacyclovir groups, respectively). Valacyclovir reduces the frequency of EBV-infected B cells when administered over a long period and, in theory, might allow eradication of EBV from the body if reinfection does not occur.

Protection from herpes simplex virus (HSV)-2 infection with replication-defective HSV-2 or glycoprotein D2 vaccines in HSV-1-seropositive and HSV-1-seronegative guinea pigs.

BACKGROUND: A herpes simplex virus (HSV)-2 candidate vaccine consisting of glycoprotein D (gD2) in alum and monophosphoryl lipid A (MPL) reduced genital herpes disease in HSV-1-seronegative women but not in men or HSV-1-seropositive women. METHODS: To determine the effect of HSV-1 serostatus on effectiveness of different vaccines, we tested gD2 in alum/MPL, gD2 in Freund's adjuvant, and dl5-29 (a replication-defective HSV-2 mutant) in HSV-1-seropositive or HSV-1-seronegative guinea pigs. RESULTS: In HSV-1-seronegative animals, dl5-29 induced the highest titers of neutralizing antibody, and after vaginal challenge with wild-type virus, dl5-29 resulted in lower rates of vaginal shedding, lower levels of HSV DNA in ganglia, and a trend for less acute and recurrent genital herpes, compared with the gD2 vaccines. In HSV-1-seropositive animals, all 3 vaccines induced similar titers of neutralizing antibodies and showed similar levels of protection against acute and recurrent genital herpes after vaginal challenge with wild-type virus, but dl5-29 reduced vaginal shedding after challenge more than did the gD2 vaccines. CONCLUSIONS: dl5-29 Is an effective vaccine in both HSV-1-seropositive and HSV-1-seronegative guinea pigs and was superior to gD2 vaccines in reducing virus shedding after challenge in both groups of animals. dl5-29 Might reduce transmission of HSV-2.

Impairment in reactivation of a latency associated transcript (LAT)-deficient HSV-2 is not solely dependent on the latent viral load or the number of CD8(+) T cells infiltrating the ganglia.

The HSV latency-associated transcript (LAT) is abundantly expressed during virus latency. Previous studies have shown that the latent viral load and CD8(+) T cells in ganglia influence the rate of reactivation of HSV. While LAT is important for efficient reactivation and establishment of latency, it is uncertain how LAT affects either the HSV latent viral load or CD8(+) T cell infiltration of ganglia. We infected mice with LAT-deficient or LAT-restored HSV-2 at a wide range of inocula. We found that the reduced rate of spontaneous ex-vivo reactivation of the LAT-deficient virus was not associated with a higher number of CD8(+) T cells in the ganglia. Reactivation rates were lower for LAT-deficient than LAT restored HSV-2 even when the latent viral loads were similar, indicating that differences in reactivation were not solely dependent on the latent viral load. Therefore, LAT likely has additional functions important for reactivation.

Serological diagnosis of human herpes simplex virus type 1 and 2 infections by luciferase immunoprecipitation system assay.

Highly quantitative and high-throughput serological tests for evaluation of humoral responses to herpes simplex virus 1 (HSV-1) and HSV-2 are not available. The efficacy of luciferase immunoprecipitation system (LIPS) assays for antibody profiling and serologic diagnosis of HSV-1 and HSV-2 infection was investigated using a panel of five recombinant HSV antigens. Plasma samples from subjects seropositive for HSV-1 and/or HSV-2 or seronegative for HSV-1 and HSV-2 that had previously been analyzed by Western blotting and the Focus Plexus immunoassay were evaluated. The LIPS test measuring anti-gG1 antibody titers was 96% sensitive and 96% specific for detecting HSV-1 infection, compared with the Focus immunoassay, and was 92% sensitive and 96% specific, compared with Western blotting. The results for the anti-gG2 LIPS test for HSV-2 precisely matched those for Western blotting, with 100% sensitivity and 100% specificity, and showed robust antibody titers in all the HSV-2-infected samples that were over 1,000 times higher than those in HSV-2-negative or HSV-1-positive samples. Antibodies to three additional HSV-2 proteins, gB, gD, and ICP8, were detected in many of the HSV-1- and/or HSV-2-infected plasma samples and showed preferentially higher immunoreactivity in HSV-2-infected plasma. The titers of antibodies to these three HSV-2 antigens also significantly correlated with each other (R=0.75 to 0.81; P<0.0001). These studies indicate that the robust anti-gG1 and anti-gG2 antibody responses detected by LIPS assays are useful for HSV-1 and HSV-2 detection and suggest that profiling of antibody responses to a panel of HSV proteins may be useful for characterizing individual humoral responses to infection and for monitoring responses to vaccines.

Comparison of immunogenicity and protective efficacy of genital herpes vaccine candidates herpes simplex virus 2 dl5-29 and dl5-29-41L in mice and guinea pigs.

A replication-defective herpes simplex virus (HSV)-2 vaccine, dl5-29, which is deleted for two essential early genes, UL5 and UL29, is highly immunogenic and protective in mice and guinea pigs. In a prior study, a derivative of HSV-2 dl5-29 termed dl5-29-41L, which has an additional deletion in UL41 (that encodes the virion-host shut-off protein), was more immunogenic and protective against challenge with wild-type HSV-2 in mice when compared with dl5-29. To determine if deletion of UL41 improves the efficacy of dl5-29 in protecting guinea pigs from HSV-2, animals were immunized with dl5-29, dl5-29-41L, or PBS. The geometric mean neutralizing antibody titers from the dl5-29 and dl5-29-41L recipients were comparable (10(1.97) and 10(2.19), respectively, p=0.15). After intravaginal challenge with wild-type HSV-2, the dl5-29-41L and dl5-29 recipients shed similar titers of HSV-2 from the vagina. Mean acute disease severity scores, numbers of recurrences during 3 months after infection, and latent viral loads in sacral ganglia were similar for dl5-29 and dl5-29-41L (all p values >0.05). dl5-29 and dl5-29-41L completely protected mice from lethal challenge with HSV-2 and induced virus-specific CD8(+) T cells in the spleens of the animals. Thus, dl5-29 was as immunogenic and protective as dl5-29-41L under these conditions. dl5-29 was at least 250,000-fold less virulent than parental virus by intracranial inoculation in healthy mice, and caused no disease in SCID mice. Both dl5-29-41L and dl5-29 are equally effective and immunogenic in guinea pigs, and dl5-29 is very safe in immunocompromised animals.

The number of herpes simplex virus-infected neurons and the number of viral genome copies per neuron correlate with the latent viral load in ganglia.

The latent viral load is the most important factor that predicts reactivation rates of animals latently infected with herpes simplex virus (HSV). To estimate the latent viral load, individual latently infected mouse trigeminal ganglia were dispersed into single cell suspensions and plated into 96-well real-time PCR plates, and HSV-2 genome copies were measured. By assuming a Poisson distribution for both the number of HSV-2 infected cells per well and the number of HSV-2 genome copies per infected cell, the numbers of infected cells and mean genome copies per infected cell were determined. Both the number of HSV-2 infected cells and the mean HSV-2 genome copy per infected cell significantly correlated with the latent viral load (p<10(-4)), indicating that both factors are responsible for the increase in the latent viral load.

Rates of reactivation of latent herpes simplex virus from mouse trigeminal ganglia ex vivo correlate directly with viral load and inversely with number of infiltrating CD8+ T cells.

Herpes simplex viruses (HSV) reactivate at rates proportional to the viral loads in latently infected ganglia. However, these rates vary substantially among infected animals. We assessed whether the numbers of HSV-specific CD8(+) T cells infiltrating latently infected ganglia also affect reactivation rates and contribute to their variability. Following corneal infection of mice with HSV type 2 (HSV-2), we quantified the latent viral loads in dissociated trigeminal ganglia by real-time PCR, the numbers of infiltrating CD8(+) T cells by flow cytometry, and the rates of reactivation by the detection of cell-free virus released from ganglion cells cultured in 96-well plates. The reactivation rates correlated directly with the latent viral loads (P = 0.001) but did so more strongly (P = 10(-7)) when cultures were depleted of CD8(+) T cells. Reactivation rates were reduced in a dose-dependent fashion by adding back ganglion CD8(+) T cells to the cultures (P = 0.003). We related the latent viral loads, numbers of CD8(+) T cells, and reactivation rates by mathematical equations. The rates of reactivation predicted from latent viral loads and numbers of infiltrating CD8(+) T cells in dissociated ganglia correlated with the observed rates of reactivation (P = 0.04). The reactivation of HSV-2 from ganglia ex vivo is determined both by the latent viral load and the number of infiltrating CD8(+) T cells.

Clonality analysis by sequence variation of the latent membrane protein 1 gene in patients with chronic active Epstein-Barr virus infection.

Chronic active Epstein-Barr virus (EBV) infection is a severe systemic disease associated with high rates of mortality and morbidity. Recent studies suggest that the clonal expansion of EBV-infected T or natural killer cells plays a crucial role in the pathogenesis of chronic active EBV infection. However, it is not clear whether chronic active EBV infection is truly a monoclonal disorder that originates from one cell. The clonality of EBV was investigated by sequence variation of the latent membrane protein 1 (LMP1) gene, which has a high degree of sequence heterogeneity. Peripheral blood mononuclear cells were obtained from nine Japanese patients with chronic active EBV infection and four with infectious mononucleosis. A carboxyl-terminal region of the LMP1 gene was analyzed by polymerase chain reaction (PCR). The amplified PCR products were subcloned, and 18 clones from each sample were sequenced. Patients with chronic active EBV infection each had two to five different LMP1 nucleotide sequences, whereas patients with infectious mononucleosis each had one to seven different sequences. Patients with chronic active EBV infection and infectious mononucleosis both had one dominant sequence. Longitudinal analysis was performed in four patients with chronic active EBV infection, in whom the dominant strains were found to have remained unchanged for several years. The results suggest that EBV in patients with chronic active EBV infection was polyclonal, although clonal expansion may occur. Collectively, these findings are critical to clarify further the pathogenesis of chronic active EBV infection and aid in the development of effective treatment strategies.

Evaluation of apoptosis in Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis.

Epstein-Barr virus (EBV) is known to be a causative agent of hemophagocytic lymphohistiocytosis (HLH). To investigate association of apoptosis in the pathogenesis of EBV-associated HLH, the serum EBV loads, and serum concentrations of soluble tumor necrosis factor receptor 1 (sTNF-R1), soluble Fas ligand, and cytochrome c were examined in 15 patients with EBV-associated HLH and 24 patients with infectious mononucleosis (IM). Levels of sTNF-R1 are known to reflect the biological activity of TNF-alpha and cytochrome c is a specific marker of apoptosis. EBV loads, and concentrations of sTNF-R1 and cytochrome c were significantly higher in patients with EBV-associated HLH than in patients with IM. On the other hand, there were no statistically significant differences in the concentrations of soluble Fas ligand. In patients with EBV-associated HLH, EBV loads, concentrations of sTNF-R1, and cytochrome c were correlated with each other. These results suggest that apoptosis, which is dependent on the EBV load and could be mediated by TNF-alpha, plays a major role in the pathophysiology of EBV-associated HLH.

Association of virus infected-T cell in severe hepatitis caused by primary Epstein-Barr virus infection.

BACKGROUND: Infectious mononucleosis owing to primary Epstein-Barr virus (EBV) infection sometimes causes hepatitis, which is usually self-limiting with mildly elevated transaminases, but can rarely develop into severe hepatitis with jaundice. OBJECTIVE: To clarify the pathogenesis of severe hepatitis by primary EBV infection. METHODS: We experienced four cases of severe hepatitis with jaundice caused by primary EBV infection. These cases were analyzed virologically and histologically, and compared with infectious mononucleosis patients without jaundice. RESULTS AND DISCUSSION: Using real-time PCR, more EBV-DNA was detected in peripheral blood from patients with severe hepatitis, as compared to those without jaundice. Furthermore, CD3+, CD4+ or CD8+ cells contained more EBV DNA than did other cell populations, indicating that in severe hepatitis, T cells harbor most of the EBV. By contrast, mainly B cells were infected in infectious mononucleosis patients without jaundice. The liver was biopsied in three of the four cases. An in situ hybridization study showed that EBV infected lymphocytes, not hepatocytes. In addition, in one patient, it was confirmed that the infected lymphocytes were CD8+ T cells. These results suggest that a large EBV burden and T cell infection may play major roles in the mechanism of severe hepatitis caused by primary EB virus infection.

Differences between T cell-type and natural killer cell-type chronic active Epstein-Barr virus infection.

Infections of T cells and natural killer (NK) cells play a central role in the pathogenesis of chronic active Epstein-Barr virus (CAEBV) infection. To characterize the virologic and cytokine profiles of T cell-type and NK cell-type infection, 39 patients with CAEBV infection were analyzed. Patients with T cell-type infection had higher titers of immunoglobulin G against early and late EBV antigens, suggesting lytic cycle infection. However, the pattern of EBV gene expression was latency type II; BZLF1, which is a hallmark of lytic cycle infection, could not be detected in any patients, regardless of infection type. Patients with CAEBV infection had high concentrations of proinflammatory, T helper cell type 1, and anti-inflammatory cytokines. The cytokine profile in patients with NK cell-type infection was similar to that in patients with T cell-type infection, but the concentration of IL-13 was high in patients with NK cell-type infection. These findings should help to clarify the pathogenesis of CAEBV infection and facilitate the development of more-effective treatments.

Comparative efficacy and immunogenicity of replication-defective, recombinant glycoprotein, and DNA vaccines for herpes simplex virus 2 infections in mice and guinea pigs.

Many candidate vaccines are effective in animal models of genital herpes simplex virus type 2 (HSV-2) infection. Among them, clinical trials showed moderate protection from genital disease with recombinant HSV-2 glycoprotein D (gD2) in alum-monophosphoryl lipid A adjuvant only in HSV women seronegative for both HSV-1 and HSV-2, encouraging development of additional vaccine options. Therefore, we undertook direct comparative studies of the prophylactic and therapeutic efficacies and immunogenicities of three different classes of candidate vaccines given in four regimens to two species of animals: recombinant gD2, a plasmid expressing gD2, and dl5-29, a replication-defective strain of HSV-2 with the essential genes UL5 and UL29 deleted. Both dl5-29 and gD2 were highly effective in attenuating acute and recurrent disease and reducing latent viral load, and both were superior to the plasmid vaccine alone or the plasmid vaccine followed by one dose of dl5-29. dl5-29 was also effective in treating established infections. Moreover, latent dl5-29 virus could not be detected by PCR in sacral ganglia from guinea pigs vaccinated intravaginally. Finally, dl5-29 was superior to gD2 in inducing higher neutralizing antibody titers and the more rapid accumulation of HSV-2-specific CD8+ T cells in trigeminal ganglia after challenge with wild-type virus. Given its efficacy, its defectiveness for latency, and its ability to induce rapid, virus-specific CD8(+)-T-cell responses, the dl5-29 vaccine may be a good candidate for early-phase human trials.

Quantitative analysis of Epstein-Barr virus (EBV)-specific CD8+ T cells in patients with chronic active EBV infection.

To clarify the pathogenesis of chronic active Epstein-Barr virus (EBV) infection, EBV-specific CD8+ T cells were enumerated, by use of human leukocyte antigen (HLA)-A*2402-restricted tetramers, in 8 patients with chronic active EBV infection, 10 patients with infectious mononucleosis, and 16 EBV-seropositive healthy control subjects. In most of the patients with chronic active EBV infection, EBV-specific CD8+ T cells were not detected. Of note, latent membrane protein 2-specific CD8+ T cells were not detectable in any patients with chronic active EBV infection. In contrast, EBV-specific CD8+ T cells were detected in patients with infectious mononucleosis and in healthy control subjects. Low frequencies of EBV-specific CD8+ T cells may be one of the immunological features of chronic active EBV infection.

Comparison of real-time and nested PCR assays for detection of herpes simplex virus DNA.

We performed a real-time PCR assay to detect herpes simplex virus (HSV) DNA, and compared it prospectively with a nested PCR assay in 164 clinical samples (109 cerebrospinal fluid and 55 sera) from patients suspected of having neonatal HSV infection or HSV encephalitis. In 25 of 164 samples, HSV DNA was detected by the nested PCR assay. All samples positive for HSV DNA in the nested PCR assay were also positive in the real-time PCR assay, and all but two samples negative for HSV DNA in the nested assay were negative in the real-time assay. The real-time PCR assay thus had a sensitivity of 100% and a specificity of 99%, when compared with the nested assay. Sequential assays in a case of disseminated HSV showed that a decrease in HSV DNA paralleled clinical improvement. Quantification of HSV DNA by real-time PCR was useful for diagnosing and monitoring patients with HSV encephalitis and neonatal HSV infection.

Smooth muscle cell outgrowth from coronary atherectomy specimens in vitro is associated with less time to restenosis and expression of a key Transcription factor KLF5/BTEB2.

Atherectomy specimens offer an opportunity to study the biology of coronary artery lesions. We cultured smooth muscle cells (SMCs) from specimens obtained from 24 patients with coronary restenosis after angioplasty to study the relationship between activity of SMCs (in vitro outgrowth) and the time course of restenosis. We also examined expression of a Kruppel-like zinc-finger transcription factor 5 (KLF; also known as BTEB2 and IKLF), which is markedly induced in activated SMCs, in the same specimens. SMC outgrowth was observed in 9 of 24 specimens (37.5%). Restenosis occurred sooner (p < 0.01) in patients whose specimens showed outgrowth compared to those whose specimens showed no outgrowth. Immunostaining for KLF5 was more common in specimens with outgrowth (89 vs. 20%, p < 0.01). These data suggest that the number of activated SMCs in lesions may determine in vitro outgrowth and also affect the time to restenosis.