Association Between Simian Virus 40 and Human Tumors

Rotondo, John Charles; Mazzoni, Elisa; Bononi, Ilaria; Tognon, Mauro; Martini, Fernanda

doi:10.3389/fonc.2019.00670

Cited by 61 publications

(67 citation statements)

References 269 publications

(411 reference statements)

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“…Each isolated DNA sample was quantified and evaluated for PCR suitability by spectrophotometric reading (NanoDrop 2000, Thermo Scientific, Monza, Italy) [ 55 ] and by amplifying the β-globin gene sequence, respectively [ 33 ]. Tight precautions were taken to avoid cross-contamination during DNA isolation procedures and PCR reactions [ 56 ]. In detail, DNA was purified simultaneously with a sample of salmon sperm DNA and a mock sample lacking DNA (distilled water) [ 57 , 58 ] and then subjected to PCR.…”

Section: Methodsmentioning

confidence: 99%

Investigation on Spontaneous Abortion and Human Papillomavirus Infection

et al. 2020

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Viral infections are considered to be risk factors for spontaneous abortion (SA). Conflicting results have been reported on the association between Human Papillomavirus (HPV) and SA. HPV DNA was investigated in matched chorionic villi tissues and peripheral blood mononuclear cells (PBMCs) from women who experienced SA (n = 80, cases) and women who underwent a voluntary interruption of pregnancy (VI; n = 80, controls) by qualitative PCR and quantitative droplet digital PCR (ddPCR). Viral genotyping was performed using real-time PCR in HPV-positive samples. Specific IgG antibodies against HPV16 were investigated in sera from SA (n = 80) and VI (n = 80) females using indirect ELISA assays. None of the DNA samples from SA subjects was HPV-positive (0/80), whilst HPV DNA was detected in 2.5% of VI women (p > 0.05), with a mean viral DNA load of 7.12 copy/cell. VI samples (n = 2) were found to be positive for the HPV45 genotype. The ddPCR assay revealed a higher number of HPV-positive samples. HPV DNA was detected in 3.7% and 5% of SA and VI chorionic tissues, respectively, with mean viral DNA loads of 0.13 copy/cell in SA and 1.79 copy/cell in VI (p >0.05) samples. All DNA samples from the PBMCs of SA and VI females tested HPV-negative by both PCR and ddPCR. The overall prevalence of serum anti-HPV16 IgG antibodies was 37.5% in SA and 30% in VI (p > 0.05) women. For the first time, HPV DNA was detected and quantitatively analyzed using ddPCR in chorionic villi tissues and PBMCs from SA and VI women. Circulating IgG antibodies against HPV16 were detected in sera from SA and VI females. Our results suggest that HPV infection in chorionic villi may be a rare event. Accordingly, it is likely that HPV has no significant role in SA.

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Section: Methodsmentioning

confidence: 99%

Investigation on Spontaneous Abortion and Human Papillomavirus Infection

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“…Quantitative real-time PCR (qPCR) has been widely employed due to (i) its use in HPV DNA load quantification, (ii) its broad detection-range of target molecules and (iii) multiplexing potential. However, qPCR has several intrinsic limitations, including: (i) low sensitivity in quantifying the amount of viral DNA when present in a low-copy number (Caraguel et al, 2011); (ii) lack of precision in estimating small differences in copy number among samples (Hindson et al, 2011); (iii) the need for calibration curves, represented by plasmid vectors carrying viral DNAs, thereby increasing the risk of false-positive results (Rotondo et al, 2019). Therefore, there has been progressive demand for more analytical assays for HPV DNA quantification.…”

Section: Introductionmentioning

confidence: 99%

Simultaneous Detection and Viral DNA Load Quantification of Different Human Papillomavirus Types in Clinical Specimens by the High Analytical Droplet Digital PCR Method

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Human papillomaviruses (HPVs) are small DNA tumor viruses that mainly infect mucosal epithelia of anogenital and upper respiratory tracts. There has been progressive demand for more analytical assays for HPV DNA quantification. A novel droplet digital PCR (ddPCR) method was developed to simultaneously detect and quantify HPV DNA from different HPV types. DdPCR was initially tested for assay sensitivity, accuracy, specificity as well as intra-and inter-run assay variation employing four recombinant plasmids containing HPV16, HPV18, HPV11, and HPV45 DNAs. The assay was extended to investigate/quantify HPV DNA in Cervical Intraepithelial Neoplasia (CIN, n = 45) specimens and human cell lines (n = 4). DdPCR and qPCR data from clinical samples were compared. The assay showed high accuracy, sensitivity and specificity, with low intra-/inter-run variations, in detecting/quantifying HPV16/18/11/45 DNAs. HPV DNA was detected in 51.1% (23/45) CIN DNA samples by ddPCR, whereas 40% (18/45) CIN tested HPV-positive by qPCR. Five CIN, tested positive by ddPCR, were found to be negative by qPCR. In CIN specimens, the mean HPV DNA loads determined by ddPCR were 3.81 copy/cell (range 0.002-51.02 copy/cell), whereas 8.04 copy/cell (range 0.003-78.73 copy/cell) by qPCR. DdPCR and qPCR concordantly detected HPV DNA in SiHa, CaSki and Hela cells, whereas HaCaT tested HPV-negative. The correlation between HPV DNA loads simultaneously detected by ddPCR/qPCR in CINs/cell lines was good (R 2 = 0.9706, p < 0.0001). Our data indicate that ddPCR is a valuable technique in quantifying HPV DNA load in CIN specimens and human cell lines, thereby improving clinical applications, such as patient management after primary diagnosis of HPV-related lesions with HPV-type specific assays.

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“…HPV DNA load values were reported as viral copies per human cell equivalents (copy/cell). Negative controls were the two samples used during the DNA extraction (i.e., salmon sperm DNA and mock samples) and two qPCR controls, including HPV-free human DNA and a non-template control [51]. Samples were run in triplicate for each qPCR assay.…”

Section: Viral Dna Load Quantificationmentioning

confidence: 99%

High Human Papillomavirus DNA loads in Inflammatory Middle Ear Diseases

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Background. Previous studies reported human papillomaviruses (HPVs) in middle ear tumors, whereas these viruses have been poorly investigated in chronic inflammatory middle ear diseases. We investigated HPVs in non-tumor middle ear diseases, including chronic otitis media (COM). Methods. COM specimens (n = 52), including chronic suppurative otitis media (CSOM) (n =38) and cholesteatoma (COMC) (n = 14), as well as normal middle ear (NME) specimens (n = 56) were analyzed. HPV sequences and DNA loads were analyzed by quantitative-PCR. HPV genotyping was performed by direct sequencing. Results. HPV DNA was detected in 23% (12/52) of COM and in 30.4% (17/56) of NME (p > 0.05). Specifically, HPV DNA sequences were found in 26.3% (10/38) of CSOM and in 14.3% (2/14) of COMC (p > 0.05). Interestingly, the HPV DNA load was higher in COMC (mean 7.47 copy/cell) than in CSOM (mean 1.02 copy/cell) and NME (mean 1.18 copy/cell) (P = 0.03 and P = 0.017 versus CSOM and NME, respectively). HPV16 and HPV18 were the main genotypes detected in COMC, CSOM and NME. Conclusions. These data suggest that HPV may infect the middle ear mucosa, whereas HPV-positive COMCs are associated with higher viral DNA loads as compared to NME.Pathogens 2020, 9, 224 2 of 10 mucosa sampled from CSOM patients [11]. However, the etiology of CSOM remains to be determined. The relationship between HPV infection and inflammation has been previously reported [12]. It has been shown that persistent infection with high-risk HPVs leads to an increase in pro-inflammatory cytokines, including IL-6, TNF-α and MIP-1α [13]. In addition, high-risk HPV type 16 (HPV16) is able to increase the expression of cyclooxygenase-2 (COX-2), a key enzyme in the synthesis of prostaglandins, which are important mediators of inflammation [14,15]. Until now, only a single study has reported HPV DNA sequences in CSOM, whereby different HPV genotypes, including HPV16, HPV18 and HPV6, have been detected in 30.7% of CSOM [4].COMC is a form of expanding growth consisting of keratinizing squamous cell epithelium [16]. There is great interest in the etiopathogenesis of HPV-associated cholesteatoma because HPV commonly infects the stratified epithelium [17,18]. However, conflicting data have been reported for HPV in COMC [10,[19][20][21]. HPV sequences have been detected in COMC with different prevalence, ranging from 3% to 70% [10,[19][20][21]. Moreover, no specific HPV genotypes have been associated with COMC, as high-and low-risk HPVs, such as HPV16, HPV18 and HPV6 and HPV11, have been detected [10,[19][20][21].There is emerging evidence that HPV infection can occur in different anatomical sites. Since HPV infects epithelia [22], all anatomical sites covered with epithelial tissue are potentially exposed to HPV infection. Apart from pluristratified tissues of the cervix [23], vulva [24] and oral pharynx [25], HPV sequences have been detected in simple epithelia from several anatomical sites such as lung [26], upper respiratory tract [27], larynx [28] and nose [29]. Since the middle ear muc...

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Association Between Simian Virus 40 and Human Tumors

Cited by 61 publications

References 269 publications

Investigation on Spontaneous Abortion and Human Papillomavirus Infection

Investigation on Spontaneous Abortion and Human Papillomavirus Infection

Simultaneous Detection and Viral DNA Load Quantification of Different Human Papillomavirus Types in Clinical Specimens by the High Analytical Droplet Digital PCR Method

High Human Papillomavirus DNA loads in Inflammatory Middle Ear Diseases

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