Positive Expression of Human Cytomegalovirus Phosphoprotein 65 in Atherosclerosis

Previous studies showed that human cytomegalovirus (HCMV) is associated with atherosclerosis. However, local vascular atherosclerosis related HCMV infection and protein expression remain unclear. This study aimed to assess the relationship between HCMV infection and atherosclerosis. Formalin-fixed, paraffin-embedded peripheral artery specimens were obtained from 15 patients with atherosclerosis undergoing vascular surgery from 2008 to 2010 at Zhongnan Hospital, Wuhan University. Pathological analyses were carried out after hematoxylin and eosin (H&E) and Masson trichrome staining. In situ hybridization and immunohistochemistry with two different monoclonal antibodies were employed to detect HCMV nucleic acids and proteins, respectively. H&E and Masson trichrome staining showed homogeneous extracellular matrix in femoral artery, while smooth muscle fibers were interlaced with collagen fibers; in carotid artery, inflammatory cell infiltration, foam cell vascular change, cholesterol crystals, and layered collagen fibers were observed. In situ hybridization showed no expression of HCMV nucleic acids in all 15 cases. Immunohistochemical staining for protein immediate-early protein (IE1 72) was negative in all cases, while phosphoprotein 65 (pp65) expression was detected in 14 cases. A high rate of positive pp65 signals was found in patients with atherosclerosis, suggesting that local HCMV infection may be associated with the pathogenesis of atherosclerosis. Further studies on this relationship are warranted.

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Detection of Human Cytomegalovirus in Patients with Epithelial Ovarian Cancer and its Impacts on Survival

10.21203/rs.2.22594/v2 ◽

2020 ◽

Author(s):

Min Yin ◽

Aiping Chen ◽

Fei Zhao ◽

Xuechao Ji ◽

Chuan Li ◽

...

Keyword(s):

Ovarian Cancer ◽

Overall Survival ◽

Epithelial Ovarian Cancer ◽

Human Cytomegalovirus ◽

Hcmv Infection ◽

Benign Tumors ◽

Early Protein ◽

Viral Carcinogenesis ◽

Formalin Fixed Paraffin Embedded

Abstract Background: The cause of epithelial ovarian cancer(EOC) is not elucidated. Viral infection may induce chronic inflammatory infection and play a role in the pathogenesis of cancers. Some viruses are considered to be oncomodulatory, modulating cellular pathways such as cell proliferation, tumor progression, vascular disease development, and immune evasion. Human cytomegalovirus (HCMV) has been detected in several types of cancers including ovarian cancer. However, the role of HCMV in ovarian carcinogenesis remains controversial. Objective: To investigate the potential role of HCMV infection in EOC, we evaluated the prevalence of HCMV proteins in EOC tissue and its impacts on patients’ survival. Methods: Formalin-fixed, paraffin-embedded tissues from 66 patients with EOC and 30 patients with benign ovarian cystadenoma were studied. Specimens were detected for expression of HCMV immediate-early protein (IE) and HCMV tegument protein (pp65) by immunohistochemistry. Results: HCMV-IE protein expression was detected in 82% of EOC and 36% of benign tumors; pp65 was detected in 97% of EOC and 63% of benign tumors. Extensive expression of HCMV-IE protein was associated with higher stage of EOC. Reactivation of latent HCMV within the tumor at interval debulking surery may be induced by neoadjuvant chemotherapy before surgery. Extensive HCMV-IE expression was associated with shorter median overall survival than focal or negative expression (39 versus 41 months, P=0.03). Multivariate analysis indicated that HCMV-IE expression was an independent prognostic factor for overall survival (P = 0.034). Conclusions: This study demonstrate a high prevalence of HCMV proteins in tissue sections from patients with EOC. HCMV infection can be potential risk factor for EOC development. Extensive HCMV-IE expression indicated a poor prognosis. The relationship between HCMV and clinical outcomes highlight the need for further researches on the oncomodulatory of HCMV in ovarian cancer. Keywords: Epithelial ovarian cancer, Human cytomegalovirus, Viral carcinogenesis, Survival

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Detection of Human Cytomegalovirus in Patients with Epithelial Ovarian Cancer and its Impacts on Survival

10.21203/rs.2.22594/v3 ◽

2020 ◽

Author(s):

Min Yin ◽

Aiping Chen ◽

Fei Zhao ◽

Xuechao Ji ◽

Chuan Li ◽

...

Keyword(s):

Ovarian Cancer ◽

Overall Survival ◽

Protein Expression ◽

Epithelial Ovarian Cancer ◽

Human Cytomegalovirus ◽

Hcmv Infection ◽

Benign Tumors ◽

Early Protein ◽

Formalin Fixed Paraffin Embedded

Abstract Background: The cause of epithelial ovarian cancer(EOC) is not elucidated. Viral infection may induce chronic inflammatory infection and play a role in the pathogenesis of cancers. Some viruses are considered to be oncomodulatory, modulating cellular pathways such as cell proliferation, tumor progression, vascular disease development, and immune evasion. Human cytomegalovirus (HCMV) has been detected in several types of cancers including ovarian cancer. However, the role of HCMV in ovarian carcinogenesis remains controversial.Objective: To investigate the potential role of HCMV infection in EOC, we evaluated the prevalence of HCMV proteins in EOC tissue and its impacts on patients’ survival.Methods: Formalin-fixed paraffin-embedded tissues from 66 patients with EOC and 30 patients with benign ovarian cystadenoma were studied. Specimens were analyzed for expression of HCMV immediate early protein (IE) and HCMV tegument protein (pp65) by immunohistochemistry. Results: HCMV-IE protein expression was detected in 82% of EOC and 36% of benign tumors; pp65 was detected in 97% of EOC and 63% of benign tumors. Extensive HCMV-IE protein expression was associated with higher stage of EOC. Reactivation of latent HCMV within the tumor at interval debulking surgery may be induced by neoadjuvant chemotherapy before surgery. Extensive HCMV-IE expression was associated with shorter median overall survival than focal or negative expression (39 versus 41 months, P=0.03). Multivariate analysis indicated that HCMV-IE expression was an independent prognostic factor for overall survival (P = 0.034). Conclusions: This study demonstrate a high prevalence of HCMV proteins in tissue sections from patients with EOC. HCMV infection can be potential risk factor for EOC development. Extensive HCMV-IE expression indicated a poor prognosis. The relationship between HCMV and clinical outcomes highlight the need for further researches on the oncomodulatory role of HCMV in ovarian cancer.

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The Expression of hsp-miRNA-200b-3p and -200c-3p in Human Cytomegalovirus-infected Formalin-Fixed, Paraffin-Embedded Tissues

Open Forum Infectious Diseases ◽

10.1093/ofid/ofx163.867 ◽

2017 ◽

Vol 4 (suppl_1) ◽

pp. S358-S358

Author(s):

Kyoung Hwa Lee ◽

Seoyeon Min ◽

Seul Gi Yoo ◽

Beom Jin Lim ◽

Jeong-Hyeon Jo ◽

...

Keyword(s):

Real Time ◽

Human Cytomegalovirus ◽

Gene Activation ◽

Invasive Disease ◽

Lytic Replication ◽

Normal Tissues ◽

Early Protein ◽

Formalin Fixed Paraffin ◽

Formalin Fixed Paraffin Embedded ◽

Formalin Fixed

Abstract Background Human cytomegalovirus (HCMV), which exist as asymptomatic latent status, can cause the tissue invasive disease through reactivation in various immunocompromised conditions. Hsp-microRNA has a specific function of post transcriptional suppression through binding with 3’ untranslated region (UTR) of mRNA. In previous study, hsp-miR-200b-3p and -200c-3p had high probability of conjugation with 3’UTR of mRNA encoded by HCMV UL 122–123 region, which translate the immediate early protein 2 (IE2) protein. IE2 (pp86) plays an essential role to initiate and regulate viral early (E) gene activation as well as propagate the subsequent steps of HCMV lytic replication. This study was aimed to evaluate whether HCMV-infected tissue had a lower expression level of hsp-miR-200b-3p and -200c-3p. Methods We had collected the formalin-fixed, paraffin-embedded tissues (FFPEs) with cytopathic pathologic findings as well as positive immunohistochemical stain (IHC) test for HCMV (N = 111). The HCMV-uninfected normal tissues (N = 77) were selected among FFPEs with neither infection nor inflammation as well as negative HCMV IHC test. We performed TaqMan® MicroRNA real-time RT-PCR to measure the expression levels of hsp-miR-200b-3p and -200c-3p and TaqMan® real-time PCR for HCMV UL83 region to measure HCMV viral load in each FFPE. We utilized the standard curves consisting of mirVanaTM miRNA mimics corresponding to each of two miRNAs, ranging from 106 to 101 copies/µL and HCMV NIBSC 09/162 strain, ranging from 5 X 106 to 5 X 101 IU/mL. Results The levels of hsp-miR-200b-3p and -200c-3p were strongly correlated with r=0.844 (P < 0.001). The expressions levels of hsp-miR-200b-3p in HCMV-infected FFPEs (log10 3.50 ± 0.13 copies/µL) were significantly lower than normal tissues (log10 5.24 ± 0.12 copies/µL of input RNA, P < 0.001). Also, HCMV-infected FFPEs were significantly lower levels of hsp-miR-200c-3p compared than normal tissues (log10 5.28 ± 0.18 vs. 7.81 ± 0.11 copies/µL of input RNA, P = 0.025). The levels of miR-200b-3p and -200c-3p had the significant inverse correlation with HCMV VL (200b-3p, spearman r=-0.392, P < 0.001 and 200c-3p, spearman r=-0.355, P < 0.001). Conclusion The low expression of hsp-miRNA-200b-3p and -200c-3p could play a pathophysiologic role of development of HCMV tissue-invasive disease. Disclosures All authors: No reported disclosures.

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Anti-IE1 CD4+ T-cell clones kill peptide-pulsed, but not human cytomegalovirus-infected, target cells

Journal of General Virology ◽

10.1099/vir.0.82958-0 ◽

2007 ◽

Vol 88 (9) ◽

pp. 2441-2449 ◽

Cited By ~ 4

Author(s):

Sandra Delmas ◽

Pierre Brousset ◽

Danièle Clément ◽

Emmanuelle Le Roy ◽

Jean-Luc Davignon

Keyword(s):

T Cells ◽

T Cell ◽

Human Cytomegalovirus ◽

Antigen Processing ◽

Hcmv Infection ◽

Target Cells ◽

T Cell Clones ◽

Early Protein ◽

Cell Clones ◽

Precise Role

Cellular immunity plays a major role in the control of human cytomegalovirus (HCMV) infection. CD4+ T lymphocytes have been shown to contribute to this function but their precise role is a matter of debate. Although CD4+ T cells have been shown to kill target cells through the perforin/granzyme pathway, whether HCMV-specific CD4+ T cells are capable of killing HCMV-infected targets has not yet been documented. In the present paper, we have taken advantage of well established cellular reagents to address this issue. Human CD4+ T-cell clones specific for the major immediate-early protein IE1 were shown to perform perforin-based cytotoxicity against peptide-pulsed targets. However, when tested on infected anitgen presenting cell targets, cytotoxicity was not detectable, although gamma interferon (IFN-γ) production was significant. Furthermore, cytotoxicity against peptide-pulsed targets was inhibited by HCMV infection, whereas IFN-γ production was not modified, suggesting that antigen processing was not altered. Remarkably, degranulation of CD4+ T cells in the presence of infected targets was significant. Together, our data suggest that impaired cytotoxicity is not due to failure to recognize infected targets but rather to a mechanism specifically related to cytotoxicity.

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Detection of Human Cytomegalovirus in Patients with Epithelial Ovarian Cancer and its Impacts on Survival

10.21203/rs.2.22594/v1 ◽

2020 ◽

Author(s):

Min Yin ◽

Aiping Chen ◽

Fei Zhao ◽

Xuechao Ji ◽

Chuan Li ◽

...

Keyword(s):

Ovarian Cancer ◽

Epithelial Ovarian Cancer ◽

Human Cytomegalovirus ◽

Benign Tumors ◽

Tissue Sections ◽

Early Protein ◽

Ovarian Carcinogenesis ◽

Formalin Fixed Paraffin ◽

Formalin Fixed Paraffin Embedded

Abstract Background: The cause of epithelial ovarian cancer(EOC) is not elucidated. It has been proved that infectious agents could contribute to ovarian carcinogenesis. Human cytomegalovirus (HCMV) has been detected in several types of tumors. Objective: To investigate the potential role of HCMV infection in EOC, we evaluated the prevalence of HCMV proteins in EOC tissue sections and its impacts on patients’ survival. Methods: Formalin-fixed, paraffin-embedded tissues from 66 patients with EOC and 30 patients with benign ovarian cystadenoma were studied. Specimens were detected for expression of HCMV immediate-early protein (IE) and HCMV tegument protein (pp65) by immunohistochemistry. Results: HCMV-IE protein expression was detected in 82% of EOC and 36% of benign tumors; pp65 was detected in 97% of EOC and 63% of benign tumors. Extensive expression of HCMV-IE protein was associated with higher stage of EOC. Reactivation of latent HCMV within the tumor at interval debulking surery may be induced by neoadjuvant chemotherapy before surgery. Extensive HCMV-IE expression was associated with shorter median overall survival than focal or negative expression (39 versus 41 months, P = 0.03). Conclusions: This study demonstrate a high prevalence of HCMV proteins in tissue sections from patients with EOC. HCMV infections can be potential risks for EOC development. The relationship between HCMV and clinical outcomes highlight the need for further researches on the oncomodulatory role of HCMV in ovarian cancer.

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Proteasome Subunits Relocalize during Human Cytomegalovirus Infection, and Proteasome Activity Is Necessary for Efficient Viral Gene Transcription

Journal of Virology ◽

10.1128/jvi.02236-09 ◽

2009 ◽

Vol 84 (6) ◽

pp. 3079-3093 ◽

Cited By ~ 44

Author(s):

Karen Tran ◽

Jeffrey A. Mahr ◽

Deborah H. Spector

Keyword(s):

Dna Replication ◽

Dna Synthesis ◽

Human Cytomegalovirus ◽

Hcmv Infection ◽

Proteasome Activity ◽

Late Gene ◽

Viral Dna ◽

Rna Transcription ◽

Viral Dna Replication ◽

Early Protein

ABSTRACT We have continued studies to further understand the role of the ubiquitin-proteasome system (UPS) in human cytomegalovirus (HCMV) infection. With specific inhibitors of the proteasome, we show that ongoing proteasome activity is necessary for facilitating the various stages of the infection. Immediate-early protein 2 expression is modestly reduced with addition of proteasome inhibitors at the onset of infection; however, both early and late gene expression are significantly delayed, even if the inhibitor is removed at 12 h postinfection. Adding the inhibitor at later times during the infection blocks the further accumulation of viral early and late gene products, the severity of which is dependent on when the proteasome is inhibited. This can be attributed primarily to a block in viral RNA transcription, although DNA synthesis is also partially inhibited. Proteasome activity and expression increase as the infection progresses, and this coincides with the relocalization of active proteasomes to the periphery of the viral DNA replication center, where there is active RNA transcription. Interestingly, one 19S subunit, Rpn2, is specifically recruited into the viral DNA replication center. The relocalization of the subunits requires viral DNA replication, but their maintenance around or within the replication center is not dependent on continued viral DNA synthesis or the proteolytic activity of the proteasome. These studies highlight the importance of the UPS at all stages of the HCMV infection and support further studies into this pathway as a potential antiviral target.

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Detection of human cytomegalovirus infection in paraffin sections by modified Steiner silver stain for LM and TEM

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100084715 ◽

1991 ◽

Vol 49 ◽

pp. 82-83

Author(s):

K. Chien ◽

Y.K. Kim ◽

I.P. Shintaku ◽

R.L. Van de Velde ◽

R.C. Heusser ◽

...

Keyword(s):

Epoxy Resin ◽

Human Cytomegalovirus ◽

Hcmv Infection ◽

Paraffin Sections ◽

Silver Stain ◽

Immunosuppressed Patients ◽

Infected Cells ◽

Methenamine Silver ◽

Methenamine Silver Stain

Human cytomegalovirus (HCMV) infection is encountered more frequently in surgical specimens due to the increased number of immunosuppressed patients (organ transplants or HIV positive). Previous studies showed that HCMV infected cells in paraffin sections can be precisely labelled by in situ hybridization or by Grocott's methenamine silver stain for LM and subsequently confirmed by TEM. However, further studies indicate HCMV infection can also be identified by modified Steiner silver stain3 but in a different manner.Modified Steiner silver stained paraffin sections can be processed for TEM as follows: Coverslips are removed by immersion in xylene. A few drops of 1% OsO4, freshly dissolved in xylene, is then pipetted onto each slide for 5 minutes. Aqueous OsO4 can not be used as it will destain the sections instantly. Slides are rinsed in xylene, xylene/acetone and then 100% acetone. The sections are infiltrated with epoxy resin/acetone, embedded in epoxy resin, polymerized and subsequently separated from slides by heat as previously described.

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Intrinsic Immune Mechanisms Restricting Human Cytomegalovirus Replication

Viruses ◽

10.3390/v13020179 ◽

2021 ◽

Vol 13 (2) ◽

pp. 179

Author(s):

Eva-Maria Schilling ◽

Myriam Scherer ◽

Thomas Stamminger

Keyword(s):

Human Cytomegalovirus ◽

Viral Entry ◽

Hcmv Infection ◽

Current Knowledge ◽

Cytidine Deaminase ◽

Regulatory Protein ◽

Detailed Knowledge ◽

Restriction Factors ◽

Immune Mechanisms ◽

Early Protein

Cellular restriction factors (RFs) act as important constitutive innate immune barriers against viruses. In 2006, the promyelocytic leukemia protein was described as the first RF against human cytomegalovirus (HCMV) infection which is antagonized by the viral immediate early protein IE1. Since then, at least 15 additional RFs against HCMV have been identified, including the chromatin regulatory protein SPOC1, the cytidine deaminase APOBEC3A and the dNTP triphosphohydrolase SAMHD1. These RFs affect distinct steps of the viral replication cycle such as viral entry, gene expression, the synthesis of progeny DNA or egress. This review summarizes our current knowledge on intrinsic immune mechanisms restricting HCMV replication as well as on the viral strategies to counteract the inhibitory effects of RFs. Detailed knowledge on the interplay between host RFs and antagonizing viral factors will be fundamental to develop new approaches to combat HCMV infection.

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E2F mediates dihydrofolate reductase promoter activation and multiprotein complex formation in human cytomegalovirus infection.

Molecular and Cellular Biology ◽

10.1128/mcb.12.10.4364 ◽

1992 ◽

Vol 12 (10) ◽

pp. 4364-4374 ◽

Cited By ~ 63

Author(s):

M Wade ◽

T F Kowalik ◽

M Mudryj ◽

E S Huang ◽

J C Azizkhan

Keyword(s):

Human Cytomegalovirus ◽

Dihydrofolate Reductase ◽

Hcmv Infection ◽

Cyclin A ◽

Immediate Early Gene ◽

Early Gene ◽

Immediate Early ◽

Gene Promoters ◽

Cellular Gene ◽

Early Protein

The adenovirus immediate-early protein E1A activates the adenovirus E2 promoter and several cellular gene promoters through transcription factor E2F. The immediate-early proteins of human cytomegalovirus (HCMV) can complement an E1A-deficient adenovirus mutant and activate the adenovirus E2 promoter. HCMV also has been shown to activate the adenovirus E2 promoter. On the basis of these findings, we have investigated whether HCMV can activate the promoter of the cellular dihydrofolate reductase (DHFR) gene, which requires E2F binding for maximal promoter activity. We show that HCMV activates the DHFR promoter and that products of the HCMV major immediate-early gene region mediate the activation of the promoter specifically through the E2F site. We used gel mobility shift assays to search for potential molecular mechanisms for this activation and found an "infection-specific" multimeric complex that bound to the E2F sites in the DHFR and E2 promoters in extracts from HCMV-infected cells but not in extracts from uninfected cells. Several antibodies against HCMV immediate-early gene products had no effect on this infection-specific complex. Subsequently, the complex was found to contain E2F, cyclin A, p33cdk2, and p107 and to be similar to S-phase-specific complexes that recently have been identified in several cell types. A functional role for the binding of the cyclin A-p33cdk2 complex to cellular gene promoters has yet to be demonstrated; however, HCMV infection causes the induction of both cellular DNA replication and transcription of growth-related genes containing E2F sites in their promoters. The findings described above therefore may relate to both of these effects of HCMV infection. We also provide evidence that some of the molecular events associated with adenovirus infection are different from those associated with HCMV infection.

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