Effect Of Herpes Simplex Virus Research Articles

ACE inhibitor, captopril, attenuates cytopathic effects of herpes simplex virus 1 in SH-SY5Y cells

Background: Over 60% of the United States population is infected with herpes simplex virus 1 (HSV-1). Current HSV-1 treatment regimens exert their antiviral effects through a common mechanism of action and suffer from high dosing frequencies, which may contribute to patient noncompliance and the subsequent development of antiviral resistance. Although primarily known for their functions in maintaining homeostatic control of arterial blood and osmotic pressures, components of the Renin-Angiotensin Aldosterone System (RAAS) have been implicated in viral replication and some components demonstrated antiviral properties. However, the antiviral properties of RAAS components have not been well characterized in reference to HSV-1. Methods: To address this gap in knowledge, we evaluated the antiviral effects of captopril, an Angiotensin Converting Enzyme 1 (ACE-1) inhibitor, on HSV-1 infection in SH-SY5Y neuroblastoma cells. We demonstrated that captopril attenuates HSV-1-induced cytopathic effects (CPE) via cell-based and morphological assays. To investigate the potential mechanism, we conducted molecular modeling studies and identified the ability of captopril to interact with and bind HSV-1 glycoprotein D. To determine where in the virus life cycle captopril exerts its protective effects, we performed experiments observing the effect of captopril on both viral entry and replication utilizing a green fluorescent protein-tagged virus and subsequent quantitative Polymerase Chain Reaction. Results: Results suggest captopril protects cells from HSV-1-induced CPE through its effect on viral replication by increasing cell viability in infected cells and decreasing virus replication, which we propose is modulated through the decreased expression of ICP0. Conclusions: Collectively, the results presented here support further evaluation of captopril, and other RAAS components, as a basis for potential novel therapeutic interventions for the treatment of HSV-1, its associated pathologies, and potentially other virus infections.

Investigating the effect of Herpes Simplex Virus 1 Latency-Associated Non-Coding RNAs on the Human Neuronal Transcriptome

Herpes simplex virus 1 (HSV-1) is a prevalent neurotropic virus that persists for the host’s lifetime due to HSV-1 establishing latency in sensory neurons. During latency, the only abundantly transcribed HSV-1 gene is the latency-associated transcript (LAT), which is processed into the 1.5kb or 2.0kb major LAT intron and several microRNAs. These latency-associated non-coding RNAs (ncRNAs) have been reported to impact the establishment, maintenance and reactivation from latency. However, the molecular mechanisms of these ncRNAs are not fully characterised, especially in the context of human neurons. This study investigated how the latency-associated ncRNAs affect the human neuronal transcriptome. We developed an experimental system to deliver the latency-associated ncRNAs to human neurons, differentiated from SH-SY5Y neuroblastoma cells. The cells were infected with a replication-defective HSV-1 mutant, in1382, that establishes a quiescent infection in which LAT is strongly expressed. Alternatively, we utilised lentiviruses engineered to express the first 3.1kb of LAT, without or with mutations in splice sites that prevents splicing of the major LAT intron, or five HSV-1 microRNAs, shown to be abundant in latently infected human ganglia. Following RNA-Seq of uninfected versus infected or transduced SH-SY5Y cells, we identified 178 host genes that had significant differential expression in response to in1382 quiescent infection and lentivirus delivery of LAT or the latency-associated microRNAs. A subset of these were validated by PCR. This work provides insight into possible roles of the latency-associated ncRNAs in neuronal cell biology and latency that could aid future investigations examining how HSV-1 latency affects human neurons.

SAT-128 An Uncommon Case of Squamous Cell Carcinoma of the Vulva with Metastasis to the Thyroid Gland

Background: Distant metastasis from Squamous Cell Carcinoma (SCC) of the Vulva is very rare and typically associated with poor outcomes. In the literature, there have been no reported cases of vulvar SCC with metastasis to the thyroid, which augments the uniqueness of the case we are presenting. Clinical Case: A 29-year-old female was hospitalized for abdominal pain & altered mental status. Labs showed calcium 21 (RR 8.5-10.5 mg/dL) with iPTH 4.3 (RR 12-88 pg/mL). Imaging revealed an 8.6 x 7.2 cm right thyroid mass (solid with cystic internal components, hyperechoic to isoechoic, wider than tall, lobulated margins, punctate echogenic foci occupying nearly the entire right lobe, minimal vascularity), mildly effacing the trachea. There were also extensive lesions consistent with systemic metastasis involving the left hilar lymph nodes, pre-tracheal lymph nodes, right hepatic lobe, head of pancreas, retroperitoneal lymph nodes, right inferior pubic ramus, proximal right humerus, left humerus, proximal femur & frontal lobe of the brain. Hypercalcemia of malignancy from an unknown cancer was diagnosed. FNA biopsy of the thyroid mass was consistent with atypia of undetermined significance. Liver biopsy showed evidence of high grade carcinoma with non-calcitonin producing neuroendocrine differentiation. Labs showed serum serotonin 11 (RR 56-244 ng/mL), CA 19-9 < 0.8 (RR 0-35 U/mL), alpha-fetoprotein 2.12 (RR 0-9 ng/mL), CEA 20.97 (RR 0-2.9 ng/mL), PTHrP 33 (RR 14-27 pg/mL), 1,25OH Vit D 18 (RR 18-72 pg/mL), chromogranin A 189 (RR 25-140 pg/mL), & calcitonin < 2 (RR < 5 pg/mL). A vaginal lesion was discovered on exam & biopsy showed squamous cells with cytopathic effect of Herpes Simplex Virus (confirmed with immunohistochemical stain). Subsequent biopsy of the brain & core needle biopsy of the thyroid showed morphology similar to a concurrent biopsy of a vulvar lesion also found on exam: poorly differentiated SCC. The patient was diagnosed with vulvar SCC with extensive metastasis. Her hospital course was complicated by atrial fibrillation, acute respiratory failure, & sepsis. She, unfortunately, passed away from her severe morbidities. Discussion: Metastasis to the thyroid is an infrequent occurrence. It is commonly encountered in breast, lung & renal cell carcinomas. It can occur due to direct spread from adjacent tissues or by lymphatic or hematogenous spread. Thyroid gland metastasis is more commonly seen in patients with aggressive or widespread carcinomas, especially by hematogenous route, due to the thyroid’s extensive vascularity. Conclusion: Thyroid gland metastasis, particularly due to vulvar SCC, is a rare entity with a poor prognosis. In patients with extensive poorly differentiated carcinoma such as our patient, it is of utmost importance to identify suspicious thyroid nodules and perform comprehensive diagnostic testing to facilitate timely intervention for improved outcomes.

Механизмы влияния вирусов простого герпеса при неразвивающейся беременности

Механизмы влияния вирусов простого герпеса при неразвивающейся беременности

The effect of herpes simplex virus vector-mediated gene therapy of protein phosphatase 1α on bladder overactivity and nociception.

We studied the effect of herpes simplex virus (HSV) vectors-based gene transfer of protein phosphatase 1α (PP1α) on bladder hypersensitivity in rats. Using adult female Sprague-Dawley rats, non-replicating HSV vectors carrying PP1α or green fluorescent protein (GFP) were injected into the bladder wall. At one week after vector inoculation, cystometry and Western blot assay were performed, whereas the other experiments were performed at 2 weeks after vector inoculation. GFP-expressing cells were identified in the bladder as well as in L6/S1 dorsal root ganglia at 14 days. In cystometry, intercontraction intervals (ICI) after resiniferatoxin (RTx; TRPV1 agonist) irrigation was significantly reduced in the PP1α group in comparison with the GFP group. Moreover, RTx-induced freezing behavior events were observed significantly more frequently in the PP1α group than the GFP group. The number of c-Fos positive cells in the L6 spinal dorsal horn was significantly less in the PP1α group than in the GFP group. Western blot assay revealed lower levels of phosphorylated inositol 1, 4, 5-triphosphate receptor (p-IP3 R), and phosphorylated TRPV1 in the PP1α compared with the GFP group. HSV vectors-mediated PP1α gene therapy may be an alternative treatment modality for cystitis-related hypersensitive bladder condition at least in part via modulation of the IP3 R signaling pathway.

Low GRK2 Underlies Hyperalgesic Priming by Glial Cell-Derived Neurotrophic Factor.

Background: We recently identified the balance between the level of G protein coupled receptor kinase 2 (GRK2) and Epac1 in nociceptors as a key factor in the transition from acute to chronic pain that occurs in mice ‘primed’ by an inflammatory stimulus. Here, we examined the contribution of GRK2 and Epac-signaling to growth factor-induced hyperalgesic priming.Methods: Mice were primed by intraplantar injection with glial cell-derived neurotrophic factor (GDNF). Mechanical allodynia in response to PGE2 was followed over time in primed and non-primed animals. GRK2 protein levels in dorsal root ganglion (DRG) neurons were quantified by immunohistochemistry. The effect of herpes simplex virus (HSV)-GRK2 amplicons to restore GRK2 levels or of an Epac inhibitor on PGE2 allodynia in primed mice was examined.Results: Glial cell-derived neurotrophic factor-induced hyperalgesia disappeared within 12 days. The hyperalgesic response to a subsequent intraplantar injection of PGE2 was prolonged from <24 h in control mice to more than 72 h in GDNF-primed mice. In male and female primed mice, PGE2 hyperalgesia was inhibited by oral administration of the Epac inhibitor ESI-09, while the drug had no effect in control mice. Mice primed with GDNF had reduced levels of GRK2 in IB4(+) small DRG neurons, but normal GRK2 levels in IB4(-) DRG neurons. Intraplantar administration of HSV-GRK2 amplicons to increase GRK2 protein levels prevented the prolongation of PGE2-induced hyperalgesia in GDNF-primed mice.Conclusion: Low GRK2 in nociceptors is critical to develop a primed state in response to GDNF and leads to engagement of Epac signaling and transition to chronic PGE2-induced hyperalgesia. Increasing GRK2 protein or inhibiting Epac signaling may represent new avenues for preventing transition to a chronic pain state.

Infection by Herpes Simplex Virus 1 Causes Near-Complete Loss of RNA Polymerase II Occupancy on the Host Cell Genome.

Lytic infection by herpes simplex virus 1 (HSV-1) triggers a change in many host cell programs as the virus strives to express its own genes and replicate. Part of this process is repression of host cell transcription by RNA polymerase II (Pol II), which also transcribes the viral genome. Here, we describe a global characterization of Pol II occupancy on the viral and host genomes in response to HSV-1 infection using chromatin immunoprecipitation followed by deep sequencing (ChIP-seq). The data reveal near-complete loss of Pol II occupancy throughout host cell mRNA genes, in both their bodies and promoter-proximal regions. Increases in Pol II occupancy of host cell genes, which would be consistent with robust transcriptional activation, were not observed. HSV-1 infection induced a more potent and widespread repression of Pol II occupancy than did heat shock, another cellular stress that widely represses transcription. Concomitant with the loss of host genome Pol II occupancy, we observed Pol II covering the HSV-1 genome, reflecting a high level of viral gene transcription. Interestingly, the positions of the peaks of Pol II occupancy at HSV-1 and host cell promoters were different. We investigated the effect of herpes simplex virus 1 (HSV-1) infection on transcription of host cell and viral genes by RNA polymerase II (Pol II). The approach we used was to determine how levels of genome-bound Pol II changed after HSV-1 infection. We found that HSV-1 caused a profound loss of Pol II occupancy across the host cell genome. Increases in Pol II occupancy were not observed, showing that no host genes were activated after infection. In contrast, Pol II occupied the entire HSV-1 genome. Moreover, the pattern of Pol II at HSV-1 genes differed from that on host cell genes, suggesting a unique mode of viral gene transcription. These studies provide new insight into how HSV-1 causes changes in the cellular program of gene expression and how the virus coopts host Pol II for its own use.

Reflectance confocal microscopy for cutaneous infections and infestations.

Reflectance confocal microscopy (RCM) is a high-resolution emerging imaging technique that allows non-invasive diagnosis of several cutaneous disorders. A systematic review of the literature on the use of RCM for the study of infections and infestations has been performed to evaluate the current use of this technique and its possible future applications in this field. RCM is particularly suitable for the identification of Sarcoptes scabies, Demodex folliculorum, Ixodes, Dermatophytes and Candida species in the clinical practice and for the follow-up after treatment. The cytopathic effect of herpes simplex virus, varicella zoster virus and molluscipoxvirus is also detectable by this imaging technique even in a pre-vesicular stage. In addition, thanks to its non-invasiveness, RCM allows pathophysiological studies.

Risk factors of renal dysfunction in patients with herpes simplex virus 2/human immunodeficiency virus-1 co-infection

Objective To explore the effects of herpes simplex virus 2 (HSV-2) co-infection on the renal function in human immunodeficiency virus-1 (HIV-1) infected patients, and to screen the risk factors of renal dysfunction. Methods A total of 302 HIV-1-infected patients were included at The Eighth People's Hospital of Guangzhou from October 2011 to February 2014, which were divided into two groups: HIV(+ )HSV(+ ) group and HIV(+ )HSV(-) group. At the same time, 143 patients with HIV(-)HSV(+ ) were included as controls. The measurements of renal function were tested and compared among groups. Pearson correlation analysis was used to explore the relationship between HSV-2 DNA level and the severity of renal dysfunction. Multivariate Logistic regression analysis was used to determine the risk factors of renal dysfunction. Results There were 128 cases in HIV-1(+ )HSV-2(+ ) group, 174 in HIV-1(+ )HSV-2(-) group and 143 in HIV-1(-)HSV-2(+ ). Compared to HIV(-)HSV(+ ) group, the level of serum creatinine (F=14.37, P<0.01), BUN (F=12.54, P<0.01), 24 h urine protein (F=16.58, P<0.01) and urine protein/creatinine ratio (F=6.37, P=0.018) in both HIV(+ )HSV(+ ) group and HIV(+ )HSV(-) group were significantly increased, while estimated glomerular filtration rate (eGFR) in these two groups were significantly decreased (F=11.96, P<0.01). Meanwhile, compared to HIV(+ )HSV(-) group, HIV-1 RNA level (t=5.876, P<0.01), serum creatinine (t=2.315, P=0.012), 24 h urine protein (t=3.648, P=0.004), and urine protein/creatinine ratio (t=2.312, P=0.012) in HIV(+ )HSV(+ ) group were significantly increased, while CD4+ T lymphocyte counts (t=4.903, P<0.01) and eGFR (t=2.275, P=0.016) were significantly decreased. Pearson correlation analysis indicated that HSV-2 DNA level was positively correlated with HIV-1 RNA level (r=0.626, P=0.002), serum creatinine level(r=0.798, P<0.01), 24 h urine protein level (r=0.702, P<0.01), and urine protein/creatinine ratio (r=0.686, P<0.01), whereas negatively correlated with CD4+ T lymphocyte counts (r=-0.796, P<0.01) and eGFR (r=-0.656, P<0.01). Multivariate Logistic regression analysis showed that HSV-2 DNA level (odds ratio [OR]=1.166, P=0.021), HIV-1 RNA level (OR=1.581, P<0.01), CD4+ T lymphocyte counts<200/μL (OR=1.762, P<0.01), age (OR=1.472, P<0.01), ≥1 complication (OR=1.062, P=0.032), and glycosylated hemoglobin level (OR=1.124, P=0.015) were independent risk factors of renal dysfunction. Conclusions HSV-2 co-infection might aggravate the renal dysfunction of HIV-1 patients, and HSV-2 DNA level might be one of the potential risk factors of renal dysfunction. Key words: HIV-1; Herpesvirus 2, human; Renal insufficiency; AIDS-associated nephropathy

Bioactive compounds from Holothuria atra of Indian ocean.

The sea cucumber (Holothuria atra) extracts have been evaluated for the presence of bioactive compounds and various biological activities. The methanol extracts showed anti proliferative activities against the Hela and MCF-7 cell lines. Similarly the inhibitory effects of Herpes simplex virus 1 and 2 cells were detected using the plaque reduction assay. The extracts of H. atra were purified using the silica gel column chromatography. The active fractions collected were observed for antimicrobial activity. The GC-MS analysis showed the availability of 59 compounds. The active bioactive compounds found in the H. atra were analyzed and their structure was identified using the 1HNMR and 13C NMR experiments.Electronic supplementary materialThe online version of this article (doi:10.1186/2193-1801-3-673) contains supplementary material, which is available to authorized users.

Differential effect of viral overexpression of nucleus accumbens shell 5-HT1B receptors on stress- and cocaine priming-induced reinstatement of cocaine seeking

5-HT1B receptors are densely expressed on terminals of medium spiny neurons projecting from the nucleus accumbens shell (NAccSh) to the ventral tegmental area, where 5-HT1B receptors modulate GABA release directly, and firing of dopaminergic neurons indirectly. While interactions between NAccSh 5-HT1B receptors and stress have been reported in early stages of psychostimulant-induced neuroadaptations, specifically psychomotor sensitization, the effect of this interaction on later stages of drug seeking is currently unknown. Here, we examined the effect of herpes simplex virus (HSV)-mediated overexpression of NAccSh 5-HT1B receptors on reinstatement of cocaine seeking induced by exposure to stress or a cocaine prime. Rats were trained to self-administer cocaine (0.75mg/kg/infusion) and the operant response was extinguished. Rats were then injected with viral vector expressing 5-HT1B and green fluorescent protein (GFP) or GFP alone into the NAccSh. The effect of 5-HT1B receptor overexpression was assessed on reinstatement induced by intermittent footshock (0.5mA for 15min) or a cocaine prime (10mg/kg, ip). Results indicate that NAccSh 5-HT1B receptor overexpression had no effect on footshock reinstatement while significantly decreasing cocaine priming-induced reinstatement. We also found that NAccSh overexpression of 5-HT1B receptors had no effect on saccharin intake following social defeat stress. These results suggest that the efficacy of pharmacological agents targeting 5-HT1B receptors for the treatment of cocaine relapse will depend largely on the nature of the reinstating stimulus. Taken together with previous results, it appears that NAccSh 5-HT1B receptors influence stress responses in early, but not in the later stages of psychostimulant-induced neuroadaptations.

Effects of Herpes Simplex Virus Vector–Mediated Enkephalin Gene Therapy on Bladder Overactivity and Nociception

We previously reported the effects of herpes simplex virus (HSV) vector-mediated enkephalin on bladder overactivity and pain. In this study, we evaluated the effects of vHPPE (E1G6-ENK), a newly engineered replication-deficient HSV vector encoding human preproenkephalin (hPPE). vHPPE or control vector was injected into the bladder wall of female rats 2 weeks prior to the following studies. A reverse-transcription PCR study showed high hPPE transgene levels in L6 dorsal root ganglia innervating the bladder in the vHPPE group. The number of freezing behaviors, which is a nociceptive reaction associated with bladder pain, was also significantly lower in the vHPPE group compared with the control group. The number of L6 spinal cord c-fos-positive cells and the urinary interleukin (IL)-1β and IL-6 levels after resiniferatoxin (RTx) administration into the bladder of the vHPPE group were significantly lower compared with those of the control vector-injected group. In continuous cystometry, the vHPPE group showed a smaller reduction in intercontraction interval after RTx administration into the bladder. This antinociceptive effect was antagonized by naloxone hydrochloride. Thus, the HSV vector vHPPE encoding hPPE demonstrated physiological improvement in visceral pain induced by bladder irritation. Gene therapy may represent a potentially useful treatment modality for bladder hypersensitive disorders such as bladder pain syndrome/interstitial cystitis.

Effects of herpes simplex virus 2 latency-associated transcript open reading frame 3 on the apoptosis in Vero cells

Objective To explore the effects of herpes simplex virus 2 (HSV-2) latency-associated transcript open reading frame 3 (LAT ORF3) gene on Vero cells against cisplatin-induced apoptosis.Methods Recombinant plasmid enhanced green fluorescent protein-open reading frame 3 (named pEGFP-ORF3) was constructed and transfected into Vero cells; then,reverse transcription (RT)-PCR was performed to detect the expression of the target gene.Cisplatin of 3 mg/L was selected to induce the apoptosis in Vero cells.Cultured Vero cells were transfected with empty plasmid and induced by cisplatin (pEGFP-C2 group),transfected with recombinant plasmid pEGFP-ORF3 and induced by cisplatin (pEGFP-ORF3 group),only induced by cisplatin (cisplatin-induced control group),or remained untreated (normal control group).Subsequently,fluorescence microscopy was conducted to observe apoptotic bodies,Giemsa stain to observe the morphology of cell nuclei,methyl thiazolyl tetrazolium (MTT) assay to evaluate cell proliferation,and flow cytometry to assess cell apoptosis.Data were assessed by using SPSS 13.0 software,and statistical analysis was carried out by one-way ANOVA and t test.Results HSV-2 333 LAT ORF3 gene was successfully cloned.The eukaryotic expression plasmid for LAT ORF3 was constructed,and the expression of LAT ORF3 gene in Vero cells was confirmed by RT-PCR.Giemsa stain showed blue-staining nuclei and pale cytoplasm in recombinant plasmid-transfected and cisplatin-induced Vero cells with a normal shape.The value of cell proliferation (absorbance at 490 nm) by MTT assay was 2.56 ± 0.21 in pEGFP-ORF3 group,similar to that in the normal control group (2.66 ± 0.13,P ＞ 0.05),but significantly higher than cisplatin-induced control group (1.65 ± 0.11,P ＜ 0.05) and pEGFP-C2 group (1.56 ± 0.18,P ＜ 0.05).As far as the apoptosis rate was concerned,no significant difference was observed between pEGFP-ORF3 group and normal control group (4.03％ ± 1.04％ vs.2.13％ ± 0.09％,P ＞ 0.05),but pEGFP-ORF3 group was statistically lower than pEGFP-C2 group (19.45％ ± 2.05％,P ＜ 0.05).Conclusion The transfected HSV-2 LAT ORF3 gene could protect Vero cells from cisplatin-induced apoptosis. Key words: Herpesvirus 2,human; Open reading frames; Latency-associated transcripts; Vero cells; Cisplatin; Apoptosis

Effect of herpes simplex virus vector-mediated interleukin-4 gene therapy on bladder overactivity and nociception

We investigated the effects of replication-defective herpes simplex virus (HSV) vector expression of interleukin-4 (IL-4) on bladder overactivity and nociception. HSV vector expressing murine interleukin-4 (S4IL4) or the control vector expressing β-galactosidase (SHZ) were injected to the rat bladder wall. At 1 week after viral injection, in cystometry performed under urethane anesthesia, the S4IL4-treated group did not show the intercontraction intervals reduction during intravesical administration of 10 nM resiniferatoxin (RTx). At 2 weeks after viral injection, behavioral studies were performed on vector-injected animals in an awakened state. Freezing behavior induced by 3 μM RTx, administered for 1 min into the bladder, was significantly suppressed in the S4IL4 group compared with the SHZ group. Murine IL-4 levels examined by ELISA were significantly increased in bladder and bladder afferent dorsal root ganglia at 2 weeks after viral injection. The expression of IL-1β and IL-2 and bladder inflammatory responses were significantly suppressed in the RTx-irritated bladder of S4IL4-injected rats. These results indicate that HSV vector-mediated interleukin-4 expression in the bladder and bladder afferent pathways reduces the inflammatory response, bladder overactivity and nociceptive behavior induced by bladder irritation in the rat model. Therefore, IL-4 gene therapy could be a new strategy for treating urinary frequency and/or bladder pain.

Reactive oxygen species drive herpes simplex virus (HSV)-1-induced proinflammatory cytokine production by murine microglia

BackgroundProduction of reactive oxygen species (ROS) and proinflammatory cytokines by microglial cells in response to viral brain infection contributes to both pathogen clearance and neuronal damage. In the present study, we examined the effect of herpes simplex virus (HSV)-1-induced, NADPH oxidase-derived ROS in activating mitogen-activated protein kinases (MAPKs) as well as driving cytokine and chemokine expression in primary murine microglia.MethodsOxidation of 2', 7'-dichlorodihydrofluorescin diacetate (H2DCFDA) was used to measure production of intracellular ROS in microglial cell cultures following viral infection. Virus-induced cytokine and chemokine mRNA and protein levels were assessed using real-time RT-PCR and ELISA, respectively. Virus-induced phosphorylation of microglial p38 and p44/42 (ERK1/2) MAPKs was visualized using Western Blot, and levels of phospho-p38 were quantified using Fast Activated Cell-based ELISA (FACE assay). Diphenyleneiodonium (DPI) and apocynin (APO), inhibitors of NADPH oxidases, were used to investigate the role of virus-induced ROS in MAPK activation and cytokine, as well as chemokine, production.ResultsLevels of intracellular ROS were found to be highly elevated in primary murine microglial cells following infection with HSV and the majority of this virus-induced ROS was blocked following DPI and APO treatment. Correspondingly, inhibition of NADPH oxidase also decreased virus-induced proinflammatory cytokine and chemokine production. In addition, microglial p38 and p44/42 MAPKs were found to be phosphorylated in response to viral infection and this activation was also blocked by inhibitors of NADPH oxidase. Finally, inhibition of either of these ROS-induced signaling pathways suppressed cytokine (TNF-α and IL-1β) production, while chemokine (CCL2 and CXCL10) induction pathways were sensitive to inhibition of p38, but not ERK1/2 MAPK.ConclusionsData presented herein demonstrate that HSV infection induces proinflammatory responses in microglia through NADPH oxidase-dependent ROS and the activation of MAPKs.

Herpes simplex viral cytopathic effect in urine cytology

Rarely are the viral cytopathic characteristic effects of herpes simplex virus (HSV) encountered in urinary cytopathology (Fig. C-1). In voided urine, such morphology is often attributed to an infection of the external genitalia. In the absence of cutaneous manifestations, however, diagnosis of HSV related pathology can suggest asymptomatic viral shedding and can potentially alert clinicians to the rarer manifestations of HSV urethritis 1,2 and HSV cystitis 3–5 . Presentations of the latter may run the gamut from an unsuspected cause of dysuria or urinary retention to life-threatening hemorrhagic cystitis in an immunocompromised host. 4,5 We present a rare finding of cellular changes consistent with HSV infection in a catheterized urine specimen collected from a 74-year-old woman with hematuria. The patient, who had a history of chronic obstructive pulmonary disease, was presented to the emergency department with dyspnea and was treated for community acquired pneumonia. Over the course of her hospitalization, she developed respiratory failure and required mechanical ventilation with steroid pulse therapy. She subsequently developed hematuria. Urine bacterial cultures were negative and microscopic examination of a catheterized urine specimen revealed frequent atypical urothelial cells amidst a background of acute inflammation and abundant erythrocytes. The abnormal cells exhibited enlarged, ground

Mechanism of DADS in the bystander effect of HSV-tk/GCV suicide gene therapy system in lens epithelial cells

To investigate the mechanism and effect of diallyl disulfide (DADS) on the bystander effect of herpes simplex virus kinase/ganciclovir (HSV-tk/GCV) suicide gene therapy system which was mediated by recombinant adeno-associated virus (rAAV) in lens epithelial cells. Immunohistochemistry method was used to determine the distribution of connexin 43 (Cx43) protein in cultured rabbit lens epithelial cells and rabbit lens epithelial cells transfected by HSV-tk suicide gene. Cx43 protein was measured and analyzed in cultured rabbit lens epithelial cells and rabbit lens epithelial cells transfected by HSV-tk suicide gene that was induced by various DADS. Cell survival was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide method. DADS increased the Cx43 protein expression in cultured rabbit lens epithelial cells and rabbit lens epithelial cells transfected by HSV-tk suicide gene, and especially in 50 μmol/L DADS. After combining with DADS, the bystander effect was significantly improved (P<0.05). DADS can elevate the expression of Cx43 protein and enhance the bystander effect of HSV-tk/GCV suicide gene therapy system.

Induction of Apoptosis by Herpes Simplex Virus-1 in Neonatal, But Not Adult, Neutrophils

We report a study on the effect of herpes simplex virus 1 (HSV-1) infection on apoptosis of neutrophils from both adults and neonates and present evidence showing that HSV-1 enhances apoptosis in neonatal, but not adult, neutrophils. HSV-1 enhanced the expression of both Fas and Fas ligand on the surface of neonatal neutrophils. Treatments with anti-Fas antibody and a Fas ligand inhibitor significantly reduced the induction of apoptosis by HSV-1. Using an ELISA assay, it was found that HSV-1 infection also leads to increased release of soluble FasL from HSV-1-infected neonatal neutrophils. Increased neonatal neutrophil apoptosis following HSV-1 infection may represent an important mechanism by which HSV-1 may diminish the antiviral response of neonatal neutrophils and might explain, at least in part, the severity of infections that are caused in newborns by this herpesvirus.

Novel therapeutic agents: Gene transfer of interleukin-4 (IL-4) produces persistent antinociception and suppresses the induction of spinal PGE2 in neuropathic pain

Inflammatory cytokines (e.g. TNF alpha, IL-1 beta and IL-6) and prostaglandin E2 in spinal cord appear to play a role in the neuropathic pain. Interleukin-4 (IL-4) is an anti-inflammatory cytokine that inhibits production of TNF alpha, IL-1 and IL-6, among other actions. In the present study, we examined the effect of herpes simplex virus (HSV)-based vector-mediated transfer of the gene coding for IL-4 (vector S4IL4) to lumbar DRG in rats with neuropathic pain. Primary DRG neurons in culture transduced with S4IL4 released IL-4 into the medium measured by ELISA. Subcutaneous inoculation of S4IL4 into the paw resulted in expression of IL-4 in L4-6 DRG and sciatic nerve in vivo detected by ELISA. Mechanical threshold was determined by stimulation of the plantar surface of the hindpaw with graded von Frey monofilaments; the threshold was established using the up-down method (Dixon). Thermal latency was assessed by thermal evoked paw withdrawal. In normal rats, neither vector expressing IL-4 nor control vector changed mechanical threshold or thermal latency. Selective L5 spinal nerve ligation (SNL) resulted in neuropathic pain characterized by mechanical allodynia and thermal hyperalgesia. Subcutaneous inoculation of vector S4IL4 produced a significant antiallodynic effect and reversed thermal hyperalgesia. Inoculation with a control HSV vector carrying the lacZ reporter transgene in the tk locus (vector SHZ) did not alter either mechanical threshold or thermal latency. Inoculation of S4IL4 one week prior to SNL delayed development of tactile allodynia and thermal hyperalgesia. Inoculation of S4IL4 suppressed gentle touch-induced expression of c-fos immunoreactivity in spinal cord. Inoculation of S4IL4 suppressed upregulation of spinal IL-1 beta and prostaglandin E2 in rats with neuropathic pain. The results demonstrate that recombinant herpes virus expressing IL-4 may be antinociceptive in the neuropathic pain.

Evaluation of the antitumor effects of Herpes simplex virus lacking ribonucleotide reductase in a murine retinoblastoma model

Purpose. To determine if an attenuated herpes simplex virus (HSV) lacking the large subunit of ribonucleotide reductase has antitumor effects in a transgenic mouse model of retinoblastoma (LHβTAg).Methods. LHβTAg mice were injected ocularly with 1 × 106 pfu of the hrR3 virus and tumor sizes were measured 3 weeks later. Replication of the virus in the eye and cultured murine retinoblastoma cells was tested by titration. Distribution of the virus in tumor was measured by X-gal staining.Results. Intraocular injection of mice with hrR3 (n = 24) did not result in a significant reduction in tumor size compared to uninjected (n = 24) or PBS injected controls (n = 16). Neither the hrR3, nor the HSV RE6 mutant, which was previously shown to have antitumor effects in vivo, replicated in cultured murine tumor cells in vitro, compared to wild-type HSV. The hrR3 virus also did not replicate significantly in tumor cells in vivo, compared to normal eye tissue.Conclusions. These results suggest that mutant HSV lacking ribonucleotide reductase do not display oncolytic activity in the LHβTAg mouse and that this model may not be suitable for studying viral oncolysis as a therapy for retinoblastoma.