Cytomegalovirus-infected Cells Research Articles

An Autopsy Case of COVID-19 with New Diffuse Pulmonary Ossification.

We present the case of a 61-year-old man who developed Coronavirus disease 2019 (COVID-19) and died during treatment for relapsing polychondritis. The patient was intubated and treated with steroid pulse therapy, remdecivir, antibacterial agents, baricitinib, and tocilizumab. However, his respiratory condition worsened, and he died 108 days after disease onset. An autopsy revealed diffuse alveolar damage in the fibrotic phase in all lung lobes, diffuse pulmonary ossification, and cytomegalovirus-infected cells in the middle lobe of the right lung. We herein discuss the clinical features and pathological findings of COVID-19 in immunosuppressed patients.

Polyploid Giant Cancer Cells Generated from Human Cytomegalovirus-Infected Prostate Epithelial Cells.

Prostate cancer is the most commonly diagnosed malignancy and the sixth leading cause of cancer death in men worldwide. Chromosomal instability (CIN) and polyploid giant cancer cells (PGCCs) have been considered predominant hallmarks of cancer. Recent clinical studies have proven the association of CIN, aneuploidy, and PGCCs with poor prognosis of prostate cancer (PCa). Evidence of HCMV transforming potential might indicate that HCMV may be involved in PCa. Herein, we underline the role of the high-risk HCMV-DB and -BL clinical strains in transforming prostate epithelial cells and assess the molecular and cellular oncogenic processes associated with PCa. Oncogenesis parallels a sustained growth of "CMV-Transformed Prostate epithelial cells" or CTP cells that highly express Myc and EZH2, forming soft agar colonies and displaying stemness as well as mesenchymal features, hence promoting EMT as well as PGCCs and a spheroid appearance. HCMV-induced Myc and EZH2 upregulation coupled with stemness and EMT traits in IE1-expressing CTP might highlight the potential role of HCMV in PCa development and encourage the use of anti-EZH2 and anti-HCMV in PCa treatment.

Polyploidy, EZH2 upregulation, and transformation in cytomegalovirus-infected human ovarian epithelial cells

Human cytomegalovirus (HCMV) infection has been implicated in epithelial ovarian cancer (OC). Polyploidy giant cancer cells (PGCCs) have been observed in high-grade serous ovarian carcinoma (HGSOC); they possess cancer stem cell-like characteristics and give rise to progeny cells expressing epithelial-mesenchymal transition (EMT) markers. EZH2 plays a potential oncogenic role, correlating with high proliferative index and tumor grade in OC. Herein, we present the experimental evidence for HCMV as a reprogramming vector that elicited human ovarian epithelial cells (OECs) transformation leading to the generation of “CMV-transformed Ovarian cells” (CTO). The infection with the two high-risk clinical strains, namely HCMV-DB and BL provoked a distinct cellular and molecular mechanisms in infected OECs. EZH2 upregulation and cellular proliferation were curtailed by using EZH2 inhibitors. The HGSOC biopsies were characterized by an elevated EZH2 expression, possessing a strong positive correlation between the aforementioned marker and HCMV. From HGSOC biopsies, we isolated three HCMV clinical strains that transformed OECs generating CTO cells which displayed proliferative potentials in addition to EZH2 upregulation and PGCCs generation; these features were reduced upon EZH2 inhibition. High-risk HCMV strains transformed OECs confirming an HCMV-induced epithelial ovarian cancer model and highlighting EZH2 tumorigenic properties. Our findings might be highly relevant in the pathophysiology of ovarian tumors thereby nominating new targeted therapeutics.

Mechanisms of Survival of Cytomegalovirus-Infected Tumor Cells.

Human cytomegalovirus (HCMV) DNA and proteins are often detected in malignant tumors, warranting studies of the role that HCMV plays in carcinogenesis and tumor progression. HCMV proteins were shown to regulate the key processes involved in tumorigenesis. While HCMV as an oncogenic factor just came into focus, its ability to promote tumor progression is generally recognized. The review discusses the viral factors and cell molecular pathways that affect the resistance of cancer cells to therapy. CMV inhibits apoptosis of tumor cells, that not only promotes tumor progression, but also reduces the sensitivity of cells to antitumor therapy. Autophagy was found to facilitate either cell survival or cell death in different tumor cells. In leukemia cells, HCMV induces a “protective” autophagy that suppresses apoptosis. Viral factors that mediate drug resistance and their interactions with key cell death pathways are necessary to further investigate in order to develop agents that can restore the tumor sensitivity to anticancer drugs.

Mechanisms of Survival of Cytomegalovirus-Infected Tumor Cells

Human cytomegalovirus (HCMV) DNA and proteins are often detected in malignant tumors, warranting studies of the role that HCMV plays in carcinogenesis and tumor progression. HCMV proteins were shown to regulate the key processes involved in tumorigenesis. While HCMV as an oncogenic factor just came into focus, its ability to promote tumor progression is generally recognized. The review discusses the viral factors and cell molecular pathways that affect the resistance of cancer cells to therapy. CMV inhibits apoptosis of tumor cells, that not only promotes tumor progression, but also reduces the sensitivity of cells to antitumor therapy. Autophagy was found to facilitate either cell survival or cell death in different tumor cells. In leukemia cells, HCMV induces a "protective" autophagy that suppresses apoptosis. Viral factors that mediate drug resistance and their interactions with key cell death pathways are necessary to further investigate in order to develop agents that can restore the tumor sensitivity to anticancer drugs.

Dynamin Inhibitors Prevent the Establishment of the Cytomegalovirus Assembly Compartment in the Early Phase of Infection.

Cytomegalovirus (CMV) infection initiates massive rearrangement of cytoplasmic organelles to generate assembly compartment (AC). The earliest events, the establishment of the preAC, are initiated in the early phase as an extensive reorganization of early endosomes (EEs), endosomal recycling compartment (ERC), trans-Golgi network (TGN), and the Golgi. Here, we demonstrate that dynamin inhibitors (Dynasore, Dyngo-4a, MiTMAB, and Dynole-34-2) block the establishment of the preAC in murine CMV (MCMV) infected cells. In this study, we extensively analyzed the effect of Dynasore on the Golgi reorganization sequence into the outer preAC. We also monitored the development of the inner preAC using a set of markers that define EEs (Rab5, Vps34, EEA1, and Hrs), the EE-ERC interface (Rab10), the ERC (Rab11, Arf6), three layers of the Golgi (GRASP65, GM130, Golgin97), and late endosomes (Lamp1). Dynasore inhibited the pericentriolar accumulation of all markers that display EE-ERC-TGN interface in the inner preAC and prevented Golgi unlinking and dislocation to the outer preAC. Furthermore, in pulse-chase experiments, we demonstrated that the presence of dynasore only during the early phase of MCMV infection (4–14 hpi) is sufficient to prevent not only AC formation but also the synthesis of late-phase proteins and virion production. Therefore, our results indicate that dynamin-2 acts as a part of the machinery required for AC generation and rearrangement of EE/ERC/Golgi membranes in the early phase of CMV infection.

Cytoplasmic virion assembly compartment of betaherpesviruses

During their life cycle, betaherpesviruses extensively reorganize the membranous system of the cell in order to develop a unique cytoplasmic environment of virion production (virus factory), known as cytoplasmic virion assembly compartment (cVAC). The betaherpesvirus cVAC has been studied both in human and murine cytomegalovirus (CMV) infected cells as they are considered to be models for betaherpesviruses biology. Studies of the cVAC were mainly based on electron microscopy and immunofluorescence tools, and in the previous decade, they have expanded to the use of new technologies and systems approaches (i.e., transcriptomics, proteomics, lipidomics), which led to the acquisition of a vast amount of data. Despite the immense expansion of knowledge about membranous organelle system, including a large amount of data derived from CMV infected cells, the organization and biogenesis of the cVAC remain unclear. Accordingly, very little is known about the processes of a final stage of CMV maturation and the virion egress. This minireview summarizes current knowledge about the biogenesis of the betaherpesvirus factory from the already existent endosomal compartments.

Pathogenic Effects and Pathogenesis Processes in Vitro & in Vivo in Murine Cytomegalovirus Infected Rat Corneal Endothelial Cells

ABSTRACT Purposes To understand the pathogenesis in rat corneal endothelial cells (RCECs) induced by murine cytomegalovirus infection in vitro and in vivo. Methods In vitro, cultured RCECs were infected with murine cytomegalovirus strain K181-eGFP (MCMV-eGFP). In vivo, experimental rats received intracameral injection of MCMV-eGFP. Replicating viruses and morphology change of RCECs in vivo were evaluated at several time points. Results In vitro, RCECs became necrosis at 6hpi. MCMV-eGFP began replicating at 12hpi. In vivo, the inflammatory reactions appeared at 12hpi, peaked at 72hpi and gradually subsided. Replicating MCMV-eGFP appeared in RCECs in vivo from 24hpi to 72hpi. RCECs enlarged after 12hpi and capsids in the nuclei were visible at 72hpi. A monocyte was found on a corneal endothelium at 120hpi. Conclusions RCECs were sensitive to MCMV in vitro. Replication of MCMV-eGFP in vivo began at 24hpi and ended after 72hpi, later than the inflammatory reactions.

Different viral doses of cytomegalovirus infection in the left and right colons

Objective: To compare the differences in the detection rates of cytomegalovirus (CMV) infection between the left and right colons for guiding endoscopic biopsy-sites. Methods: The cases of colonic CMV infection confirmed by histology at the Peking Union Medical College Hospital from July 2015 to July 2018 were collected and analyzed. Splenic curvature was used as the dividing point between the left and right colons. The CMV inclusions were identified histologically, and the CMV infected cells were detected using immunohistochemistry (EnVision method). The numbers of infected cells in the left and right colons were compared by histological and immunohistochemical results. A total of 731 patients were identified. Ten of the 731 patients had histological specimens of both left and right colons and were confirmed with CMV EnVision immunohistochemical study. There were 25 biopsy or resection specimens. Seven patients were male and 3 were female, and their ages ranged from 29 to 66 years, with a median age of 55 years. All of the 10 patients were also diagnosed with ulcerative colitis. Results: The number of the cells infected with CMV in the left colon was 115 (1-41), while that in the right colon was 76 (0-51). In 8/10 cases, the number of CMV infected cells in the left colon was more than that in the right colon. Conclusions: The study on the biopsies and resection specimens suggests that CMV infection is mostly present in the left colon. The left colon thus might be an important endoscopic biopsy-site for the cases with suspicion of CMV infection.

Human Herpesvirus 6 as an Indicator of Cytomegalovirus Infection and Its Attributable Disease Symptoms in Liver Transplant Recipients.

Human β-herpes viruses, including cytomegalovirus and human herpesvirus 6, can become activated in liver transplant patients. Here, we evaluated the effects of human herpesvirus 6 infection as an independent factor on cytomegalovirus infection and the occurrence of associated diseases in liver transplant patients. In this cross-sectional study, 46 patients who underwent deceased-donor liver transplant at Nemazi Hospital, Shiraz University of Medical Sciences (Shiraz, Iran) were prospectively monitored for cytomegalovirus and human herpesvirus 6 infections during 3 months posttransplant. Taq-man real-time polymerase chain reaction assay as an accurate and rapid test and pp65-anigenemia as the standard test were used to monitor cytomegalovirus infections, whereas human herpesvirus 6 infection was monitored by Taq-man real-time polymerase chain reaction assay. We also followed clinical findings from laboratory data and symptoms of cytomegalovirus infections. Active cytomegalovirus infection was detected in 23 liver transplant recipients (50%), of which 17 (74%) were diagnosed with cytomegalovirus-related diseases. Active human herpesvirus 6 infection was detected in 25 patients (54%). Thirteen of 17 cytomegalovirussymptomatic patients had coinfection with human herpesvirus 6. In 10 ofthe 13 patients with coinfection, human herpesvirus 6 DNAemia appeared significantly earlier by 9 days than cytomegalovirus infection. In the pp65 antigenemia test, the mean number of cytomegalovirus-infected polymorphonuclear cells was 42.47 ± 5.41, which was correlated with incidence of clinical presentation. In symptomatic patients, average serum and polymorphonuclear cell viral loads of cytomegalovirus were 12064.59 copies/mL and 6735 copies/2 × 105 cells,respectively, with significant differences between the loads and cytomegalovirusattributable disease symptoms. Average human herpesvirus 6 DNA burden in serum samples of symptomatic patients was 11283 copies /mL, which was statistically related to cytomegalovirus-attributable disease symptoms. We found that human herpesvirus 6 infection is often associated with cytomegalovirus reactivation and cytomegalovirus-attributable disease symptoms.

P2Y2 purinergic receptor modulates virus yield, calcium homeostasis, and cell motility in human cytomegalovirus-infected cells

Human cytomegalovirus (HCMV) manipulates many aspects of host cell biology to create an intracellular milieu optimally supportive of its replication and spread. Our study reveals that levels of several components of the purinergic signaling system, including the P2Y2 and P2X5 receptors, are elevated in HCMV-infected fibroblasts. Knockdown and drug treatment experiments demonstrated that P2Y2 enhances the yield of virus, whereas P2X5 reduces HCMV production. The HCMV IE1 protein induces P2Y2 expression; and P2Y2-mediated signaling is important for efficient HCMV gene expression, DNA synthesis, and the production of infectious HCMV progeny. P2Y2 cooperates with the viral UL37x1 protein to regulate cystolic Ca2+ levels. P2Y2 also regulates PI3K/Akt signaling and infected cell motility. Thus, P2Y2 functions at multiple points within the viral replication cycle to support the efficient production of HCMV progeny, and it may facilitate in vivo viral spread through its role in cell migration.

Cytomegalovirus hepatitis after bone marrow transplantation: An autopsy study with clinical, histologic and laboratory correlates.

Mortality from cytomegalovirus disease after marrow transplantation can be reduced by treatment with antiviral drugs based on the detection of viremia and organ involvement. We examined autopsy liver specimens to determine the frequency, extent and outcome of cytomegalovirus hepatitis and whether cytomegalovirus hepatitis occurred in the absence of cytomegalovirus disease elsewhere. Autopsy specimens from 50 transplant patients were evaluated for cytomegalovirus-infected cells, in five groups of 10, according to extent of CMV during life and at autopsy. Liver sections were examined by routine light microscopy, immunohistochemistry and in situ DNA hybridization. Clinical and laboratory data collected during the last 30days of life were analysed as markers of liver cytomegalovirus infection. Cytomegalovirus-infected cells were detected in the livers of 10/10 patients with cytomegalovirus infection during life and widespread cytomegalovirus at autopsy; in 3/20 livers from patients with cytomegalovirus infection during life but negative liver cultures at autopsy; and in 1/10 livers from cytomegalovirus-seropositive patients who had been without other evidence of cytomegalovirus infection. Histology detected a lower density of cytomegalovirus-bearing cells per unit area of liver, compared to immunohistochemistry and in situ hybridization. No cytomegalovirus-infected cells were detected in livers from cytomegalovirus-seronegative controls. No distinctive clinical or laboratory findings correlated with liver cytomegalovirus detection. CMV liver disease is common in allografted patients with disseminated CMV but may rarely be isolated to the liver, best demonstrated with IHC and ISH. Massive hepatic necrosis from CMV was not seen in any autopsy liver in this study.

Abstract 614: Cytomegalovirus (CMV) reactivation and patient survival in head and neck cancer

Abstract There is building evidence that cytomegalovirus (CMV) may contribute to the development of solid tumors, however to date this has not been investigated in head and neck cancer (HNC). CMV is a highly prevalent, latent herpesvirus infecting over 50% of the population. CMV maintains latency in CD34+ cells, and recent work has demonstrated these latently infected cells preferentially differentiate into immunosuppressive monocytes. However, CMV is not solely immune suppressive as highly cytotoxic adaptive natural killer (NK) cells arise specifically in response to CMV reactivation. Here, we have investigated whether biomarkers of CMV reactivation are associated with patient survival in non-smoking HNC (n=184). CMV DNAemia was assessed using digital PCR, and a threshold of 1000 copies/mL serum was defined as reactivation; 22% of patients met this definition of CMV reactivation, however they did not have significantly worse survival than those without CMV reactivation (HR 1.5 (95% CI 0.7 - 3.0)). Monocytes that mobilize to the cancer site contribute to tumor-associated inflammation. Therefore, we explored whether a biomarker of inflammation, CRP, modified the association of CMV reactivation and HNC patient survival. In the absence of inflammation, there was no association between CMV reactivation and patient survival (HR 0.90 (95% CI 0.30 - 2.71)). However, those with both CMV reactivation and high inflammation had dramatically worse survival (HR 8.86 (95% CI 3.63 - 21.64)). These data are consistent with the recent report that CMV infected monocytes are immunosuppressive and suggest that stimulation of these monocytes to the tumor site may negatively impact patient survival. Future work should address the generalizability of this finding to other cancer types and determine whether CMV infected myeloid cells are present in the tumor microenvironment. Citation Format: Heather H. Nelson, Jeffrey S. Miller, DeVon Hunter-Schlichting, Rondi A. Butler, Karl T. Kelsey. Cytomegalovirus (CMV) reactivation and patient survival in head and neck cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 614.

The Membrane-Spanning Peptide and Acidic Cluster Dileucine Sorting Motif of UL138 Are Required To Downregulate MRP1 Drug Transporter Function in Human Cytomegalovirus-Infected Cells.

The human cytomegalovirus (HCMV) UL138 protein downregulates the cell surface expression of the multidrug resistance-associated protein 1 (MRP1) transporter. We examined the genetic requirements within UL138 for MRP1 downregulation. We determined that the acidic cluster dileucine motif is essential for UL138-mediated downregulation of MRP1 steady-state levels and inhibition of MRP1 efflux activity. We also discovered that the naturally occurring UL138 protein isoforms, the full-length long isoform of UL138 and a short isoform missing the N-terminal membrane-spanning domain, have different abilities to inhibit MRP1 function. Cells expressing the long isoform of UL138 show decreased MRP1 steady-state levels and fail to efflux an MRP1 substrate. Cells expressing the short isoform of UL138 also show decreased MRP1 levels, but the magnitude of the decrease is not the same, and they continue to efficiently efflux an MRP1 substrate. Thus, the membrane-spanning domain, while dispensable for a UL138-mediated decrease in MRP1 protein levels, is necessary for a functional inhibition of MRP1 activity. Our work defines the genetic requirements for UL138-mediated MRP1 downregulation and anticipates the possible evolution of viral escape mutants during the use of therapies targeting this function of UL138.IMPORTANCE HCMV UL138 curtails the activity of the MRP1 drug transporter by reducing its steady-state levels, leaving cells susceptible to killing by cytotoxic agents normally exported by MRP1. It has been suggested in the literature that capitalizing on this UL138-induced vulnerability could be a potential antiviral strategy against virally infected cells, particularly those harboring a latent infection during which UL138 is one of the few viral proteins expressed. Therefore, identifying the regions of UL138 required for MRP1 downregulation and predicting genetic variants that may be selected upon UL138-targeted chemotherapy are important ventures. Here we present the first structure-function examination of UL138 activity and determine that its transmembrane domain and acidic cluster dileucine Golgi sorting motif are required for functional MRP1 downregulation.

A gB/CD3 bispecific BiTE antibody construct for targeting Human Cytomegalovirus-infected cells

Bispecific T cell engager (BiTE) antibody constructs are successfully used as cancer therapeutics. We hypothesized that this treatment strategy could also be applicable for therapy of human cytomegalovirus (HCMV) infection, since HCMV-encoded proteins are abundantly expressed on the surface of infected cells. Here we show that a BiTE antibody construct directed against HCMV glycoprotein B (gB) and CD3 efficiently triggers T cells to secrete IFN-γ and TNF upon co-culture with fibroblasts infected with HCMV strain AD169, Towne or Toledo. Titration of gB expression levels in non-infected cells confirmed that already low levels of gB are sufficient for efficient triggering of T cells in presence of the BiTE antibody construct. Comparison of redirecting T cells with the bispecific antibody versus a chimeric antigen receptor (CAR) based on the same scFv showed a similar sensitivity for gB expression. Although lysis of infected target cells was absent, the BiTE antibody construct inhibited HCMV replication by triggering cytokine production. Notably, even strongly diluted supernatants of the activated T cells efficiently blocked the replication of HCMV in infected primary fibroblasts. In summary, our data prove the functionality of the first BiTE antibody construct targeting an HCMV-encoded glycoprotein for inhibiting HCMV replication in infected cells.

A Rare Case of Cytomegalovirus Colitis in an Immunocompetent Patient Mimicking Clostridium difficile Colitis

Cytomegalovirus (CMV) colitis usually occurs in immunocompromised patients and can cause significant morbidity and mortality. We report a 68 year old immunocompetent male presenting with abdominal pain and diarrhea who was found to have CMV colitis. 68 year old african american male with a history of end stage renal disease on hemodialysis presented with a one month history of watery diarrhea, diffuse abdominal pain, and weakness. Patient reported a 15-lb weight loss. No blood or mucus seen in the stool. He denied nausea or vomiting, dietary changes, international travel, sick contacts, or recent antibiotic use. Physical examination revealed tachycardia, dry mucous membranes, and a normal abdominal exam. Laboratory data was only remarkable for electrolyte abnormalities: sodium 150 mg/dL, potassium 2.1 mg/dL, and phosphorus 1.1 mg/dL. The patient received intravenous fluids and his electrolytes were corrected. CBC, TSH, urinary 5-HIAA, and celiac panel were normal. Fecal leukocytes, clostridium difficile toxin PCR, rotavirus ELISA, bacterial cultures, giardia, and cryptosporidium antigen were negative. An abdominal computed tomography was concerning for an infectious or inflammatory colitis in the descending and sigmoid colon. ESR, CRP, and fecal calprotectin were elevated. Patient underwent a colonoscopy with biopsies, which revealed diffuse viability with ulcers, exudates and pseudomembranes concerning for clostridium difficile (CD) colitis. Patient was started on oral vancomycin. Despite treatment, his diarrhea did not improve. Later, colonic biopsies surprisingly revealed cytopathic evidence consistent with CMV colitis ( Figure 1 & 2). CMV viral load was elevated at 2592 copies/mL. Human immunodeficiency virus serology was negative. Vancomycin was stopped, and the patient was started on ganciclovir with resolution of diarrhea.1554_A Figure 1. Left image: the arrows indicate CMV “owl's eye inclusions on H & E ( 40x) Right image: corresponding CMV infected cells light up brown with IHC (immunohistochemistry) stain. (40x)1554_B Figure 2. Left Image: the arrows indicate CMV “owl's eye inclusions on H & E ( 20 x) Right Image: corresponding CMV infected cells light up brown with IHC (immunohistochemistry) stain. ( 20 x)CMV colitis rarely occurs in immunocompetent patients making it a diagnostic challenge. Furthermore, it can mimic CD colitis because of some similar overlapping endoscopic findings. Recognizing that CMV colitis may occur in immunocompetent patients can prevent serious morbidity and mortality, especially in those with advanced age and multiple medical comorbidities.

Human cytomegalovirus-infected cells release extracellular vesicles that carry viral surface proteins

Extracellular vesicles (EVs) released by virus-infected cells typically incorporate host and viral components inside the vesicles (cargo molecules). Here, we investigated if human cytomegalovirus (HCMV) proteins are incorporated in EV outer membrane released by HCMV-infected cells. We separated EVs from HCMV using an iodixanol step-gradient and found that the separated vesicles carried EV markers such as the tetraspanin CD63 and Rab27A. Flow analysis of individual EVs demonstrated that on average, 15 ± 3.7% of EVs were positive for gB, 5.3 ± 2.3% were positive for gH and 3.74 ± 1.5% were positive for both gB and gH. In light of previous findings demonstrating HIV envelope proteins in EV membranes, the presence of viral protein at the surface of EVs released by HCMV-infected cells indicated that viral membrane proteins incorporated in EVs released by virus-infected cells may be a general phenomenon.

Characterization of phenyl pyrimidine derivatives that inhibit cytomegalovirus immediate-early gene expression.

BackgroundPreviously, we established a reporter cell line for human cytomegalovirus and screened anti-human cytomegalovirus compounds using the cell line. In this study, we characterized one of the identified compounds, 2,4-diamino-6–(4-methoxyphenyl)pyrimidine (coded as 35C10).Methods50% Effective concentrations (EC50s) and 50% cytotoxic concentrations (CC50s) of 35C10 and its derivatives in human fibroblasts were determined by X-gal staining of the cells infected with human cytomegalovirus Towne strain expressing β-galactosidase.ResultsEC50 and CC50 of 35C10 were 4.3 µM and >200 µM, respectively. Among several 35C10 derivatives, only one lacking 4-amino group of pyrimidine showed a similar EC50. 35C10 weakly inhibited murine cytomegalovirus, herpes simplex virus type 1, and varicella-zoster virus. A “time of addition” experiment suggested that 35C10 inhibited an early phase of the infection. Although 35C10 did not inhibit viral attachment to the cells nor the delivery of viral DNA to the nuclei, it decreased the number of infected cells expressing immediate-early 1 and 2 (IE1/IE2) proteins. 35C10 also inhibited the activation of a promoter for TRL4 in the reporter cells upon human cytomegalovirus infection, but not in the same reporter cells transfected with a plasmid expressing IE2.ConclusionOur findings suggest that 35C10 is a novel compound that inhibits IE gene expression in human cytomegalovirus-infected cells.

Interaction of Immunoglobulin with Cytomegalovirus-Infected Cells.

Intravenous immunoglobulin (IVIG) is used to treat or prevent severe viral infection, especially cytomegalovirus (CMV) infections. IVIG was characterized to understand its interaction with CMV-infected cells. IVIG retarded CMV spread and reduced virus yields depending on the neutralizing (NT) antibody titer. Immediate early protein synthesis was reduced by IVIG in 3 to 15 h, and IVIG specifically reduced the ratio of 66/68k protein synthesis among immediate early proteins in an NT antibody-dependent manner between 4 and 8 h after infection, indicating that antigenic modulation of CMV-infected cells by IVIG reduced viral protein synthesis and virus production. The half-life of antibody bound to CMV-infected cells was 3.8 h. NT antibody titers to varicella-zoster virus (VZV) and CMV in IVIG were dose dependently absorbed by cells infected with VZV and CMV, respectively, but the antibody titers to CMV and VZV, respectively, were not affected. NT antibody in 0.3 mL of IVIG (15 mg) was specifically absorbed by 108 CMV-infected cells and 107 VZV-infected cells, suggesting that the NT antibody in IVIG might be inactivated by one-tenth of a similar volume of CMV-infected or VZV-infected cells. Various antiviral activities of IVIG may contribute to control and alleviation of CMV infection.

MiR-138 promotes migration and tube formation of human cytomegalovirus-infected endothelial cells through the SIRT1/p-STAT3 pathway.

Human cytomegalovirus (HCMV) has been reported to be linked to vascular disease through the induction of neovessel formation. We have previously reported that microRNA (miR)-217 and miR-199a-5p enhance endothelial angiogenesis via inhibition of sirtuin 1 (SIRT1) in HCMV-infected human umbilical vein endothelial cells (HUVECs). Here, we found that miR-138 also suppressed the expression of the SIRT1 protein and stimulated phosphorylation of signal transducer and activator of transcription 3 (p-STAT3). Moreover, the regulation of p-STAT3 expression mediated by SIRT1 was found to promote HCMV-induced angiogenesis. These findings revealed that miR-138 might promote angiogenesis of HCMV-infected HUVECs by activating the SIRT1-mediated p-STAT3 pathway, and this could provide novel insights into HCMV-induced angiogenesis.