Detection of human papillomavirus DNA in laryngeal squamous cell carcinoma by polymerase chain reaction

Although human papillomaviruses (HPVs) have been found in many, but not all, tumours of the oral cavity, nose, pharynx and larynx, the true role of HPV in malignant tumours of the head and neck is still unclear. The presence of HPV DNA was investigated in 45 fresh squamous cell carcinoma (SCC) specimens and in 29 normal mucosa specimens collected from 45 primary laryngeal SCC patients. HPV DNA was detected using the polymerase chain reaction (PCR) with consensus primers that detect HPV types 6, 11, 16 and 18.9 of the 45 patients (20%) were HPV positive; the presence of HPV was also detected in the corresponding normal laryngeal mucosa of four of the 29 specimens (14%). No statistically significant differences were found between the presence of HPV DNA in normal specimens and in neoplastic mucosa specimens. No correlation was found between HPV DNA positive tumours and size, T classification, lymph node involvement and histological grading. This study adds further evidence suggesting a possible role of HPV DNA infection in laryngeal carcinogenesis.     

Comparison of human immunodeficiency virus 1 DNA polymerase chain reaction and qualitative and quantitative RNA polymerase chain reaction in human immunodeficiency virus 1-exposed infants

The endothelial lining of the blood-brain barrier tightly controls the distribution of peptide hormones between the central nervous system and the circulation. By using primary cultures of brain microvessel endothelial cells, an in vitro model of the blood-brain barrier, we report here the uptake and transport of the octapeptide angiotensin II by a specific receptor population. With the angiotensin II antagonists losartan (AT1 specific) and PD 123,319 (AT2 specific), we showed that both the uptake and transport of angiotensin II were mediated by the AT1 receptor. Western blot analysis confirmed the existence of the AT1 receptor in our cell-culture model. Rhodamine 123 studies also suggested that both angiotensin II antagonists, but not angiotensin II, were substrates for the P-glycoprotein efflux system, thus restricting the transport of these compounds. These results suggest an AT1 receptor mediates uptake and transport of angiotensin II at the blood-brain barrier and may contribute to the regulation of cerebrovascular levels of the peptide.     

Further study of hepatitis C virus RNA detection in formalin-fixed, paraffin-embedded liver tissues by ligation-dependent polymerase chain reaction

AIM: Refractive cataract surgery using corneal incisions is aiming at neutralization of preoperative astigmatism. PATIENTS AND METHODS: 61 patients with preoperative astigmatism of 2.25 +/- 0.98 were included in the treatment. A self-sealing corneal tunnel incision measuring 4.0 to 4.1 mm in external diameter and 6.5 to 7.0 mm in internal diameter (stretch incision) was performed on the steeper axis. After capsulorhexis and phacoemulsification a 5 mm PMMA lens was implanted without suturing. Keratometry and corneal topography were performed preoperatively, 3 days and 1 year respectively following surgery. The statistical analysis was based on the Wilcoxon signed ranks test. RESULTS: Surgical induced astigmatism (IA) following superior incisions in cases of astigmatism with the rule (n = 29) amounted to 1.93 +/- 0.97, while lateral incisions in cases of astigmatism against the rule (n = 29) led to an IA of 1.35 +/- 0.73. Axial shifts by more than 30 degrees were 23% following superior incisions and 17%, after lateral incisions. We observed. astigmatic reduction of 1.3 D after superior incisions and 0.7 D following lateral incisions. CONCLUSION: By 4 mm corneal cataract incisions on the steeper axis a high preoperative astigmatism can be reduced significantly without additional keratotomies.     

Detection of human papillomavirus gene sequences in laryngeal tumors and premalignant changes by polymerase chain reaction

Human papillomavirus gene sequences have been detected in a number of malignant and benign tumours. Non-oncogenic types 6 and 11 are etiological factors of benign mucosal tumours. Types 16 and 18 can be detected in malignancies most often but their role in the etiopathogenesis of cancers is still unclear. In our study we examined formalin-fixed and paraffin-embedded archive laryngeal tissues containing squamous cell carcinoma, papilloma and precancerous lesions for the presence of human papillomavirus genes. As a control we also examined tissues harbouring laryngeal nodules which represented the normal larynx in our study. After DNA preparation from the paraffin blocks we performed polymerase chain reaction to detect the DNA of human papillomavirus types 6, 11, 16 and 18. In the squamous cell carcinomas, papillomas and precancerous lesions the presence of human papillomavirus gene sequences was significantly higher than in the control group. To verify the integrity of DNA we also amplified a sequence deriving from the cellular beta-globin gene. Based on the 100% positivity for this gene we declare that the combination of our DNA preparation and polymerase chain reaction is a reliable method for detecting DNA sequences from formalin-fixed and paraffin-embedded tissues.     

Detection of EBV in tumor tissue and peripheral blood by polymerase chain reaction in patients with nasopharyngeal carcinoma

Sixty-nine untreated patients with a pathologically verified nasopharyngeal carcinoma were selected for the study of detection of EBV in nasopharyngeal tumor and peripheral blood by polymerase chain reaction (PCR). Primers were directed to conserved regions of EBV genome encoding capsid protein gp 220 (Bam HI L region). A distinct 239 bp band of the PCR products indicated the presence of EBV. Results showed that EBV DNA was obtained in 91.3% of 69 NPC patients and 16.7% of 18 healthy individuals on nasopharyngeal tissue, and the difference was statistically significant between the above two groups. Nevertheless, no EBV DNA was verified from the mononuclear cells of the peripheral blood of the two groups. There was no relationship between the positive EBV DNA and the titer of serological markers. Meanwhile, the positive EBV DNA did not show any relationship with the histology type, tumor and nodal bulk, or even metastasis. Although a high positive rate of EBV DNA was detected in nasopharyngeal tumor of patients, additional environmental and genetic factors must still be considered.     

Human papillomavirus type 16 DNA detected by the polymerase chain reaction in non-cancer tissues of the head and neck

Cancer-free tissues from various anatomical subsites in the head and neck were examined by the polymerase chain reaction (PCR) for the incidence of human papillomavirus (HPV) types 16 and 18. We detected HPV-16 DNA in 9 of 103 samples (8.7%), including specimens from the paranasal sinuses, tonsil, hypopharynx and larynx. However, no HPV-16/18 DNA was detected by Southern hybridization in these 9 samples. The significance of the presence of HPV-16 DNA in non-cancer tissues is still unknown, but PCR detection only of high-risk HPV DNA in head and neck cancer should be evaluated cautiously because of its ubiquity in this region.     

The presence of human papillomavirus in sinonasal papillomas, demonstrated by polymerase chain reaction with consensus primers

The frequency of human papillomavirus (HPV) in sinonasal papillomas seems to vary considerably. The highest frequencies have been reported by investigators using in situ DNA or RNA hybridization. Few studies have used polymerase chain reaction, and in these reports the frequency of HPV detection is rather low. We have investigated the presence of HPV in sinonasal papillomas using the polymerase chain reaction with a set of degenerated consensus primers, which amplify the vast majority of the known HPV types. Human papillomavirus was found in three of 14 papillomas. By in situ hybridization the same three papillomas were positive for HPV type 6/11.     

The prevalence of herpes family virus DNA in the conjunctiva of patients positive and negative for human immunodeficiency virus using the polymerase chain reaction

OBJECTIVE: To help understand the pathogenesis of herpes family virus ocular infection among patients positive for HIV, the authors compared the rates of detection of herpes family virus DNA from the conjunctiva of patients who are positive and negative for human immunodeficiency virus (HIV) using the polymerase chain reaction (PCR). DESIGN: Cross-sectional study. PARTICIPANTS: The conjunctival scrapings of 30 patients positive for HIV and 30 patients negative for HIV were examined. INTERVENTION: PCR was used to assay for the presence of herpes simplex virus type 1 (HSV), varicella-zoster virus (VZV), cytomegalovirus (CMV), and Epstein-Barr virus (EBV) DNA (n = 240 samples). MAIN OUTCOME MEASURE: The rate of detection of virus DNA in the two groups, controlling for age, gender, and race, was measured. RESULTS: HSV and VZV DNA were not detected in any of the HIV-positive or HIV-negative samples. CMV DNA was detected in 20% (6 of 30) of patients positive for HIV and was undetected in control subjects negative for HIV (P = 0.01). EBV DNA was detected in 40% (12 of 30) of patients positive for HIV and in 47% (14 of 30) of control subjects negative for HIV (P = 0.58). CONCLUSIONS: There was no difference in the frequency of detection of HSV, VZV, or EBV DNA from the conjunctiva of patients positive or negative for HIV. Only CMV DNA was detected at a significantly higher rate in the conjunctiva of patients positive for HIV compared with control subjects negative for HIV. These different rates of peripheral virus shedding may be one possible explanation for the different rates of clinical infection among the herpes family viruses among patients positive for HIV.     

Detection of human papillomavirus in cervical scrapings by in situ hybridization and the polymerase chain reaction in relation to cytology

A gynaecological out-patient population consisting of 200 patients aged 19-43 years (mean age 34.2 years) was screened for the presence of human papillomavirus (HPV) by the polymerase chain reaction and in situ hybridization on cervical scrapings. A novel method was applied for the detection of HPV in cervical cells by embedding them in a paraffin block before in situ hybridization was performed. This technique resulted in well preserved cytological morphology, easy performance and economy of probes. In eight of the 200 patients (4%), human papillomavirus DNA was revealed by the polymerase chain reaction. Subtyping revealed the presence of HPV serotype 16 DNA in three of these patients. In one patient HPV serotype 18 DNA was also present. The in situ hybridization assay was able to detect all those cases with a specific HPV serotype infection.     

Prevalence of human papillomavirus in squamous cell carcinoma of the head and neck determined by polymerase chain reaction and Southern blot hybridization: proposal for optimized diagnostic requirements

OBJECTIVE: To develop Canadian projections for the prevalence and numbers of people with arthritis and arthritis disability, overall and in major age groups. METHODS: Age and sex specific data from the 1991 General Social Survey and the 1994 National Population Health Survey on the prevalence of arthritis and arthritis disability were applied to population projections for Canada for every 5 years between 1991 and 2031. RESULTS: Between 1991 and 2031 we project that the prevalence of arthritis diagnosed by a health professional as a longterm condition in Canada will increase from 10.7 to 15.7%, an increase of 46.7%, and the number of people with arthritis will increase from 2.9 to 6.5 million, an increase of 124%. Comparable changes in prevalence and numbers of people with self-reported arthritis are 17.1% (4.7 million) to 23.6% (9.7 million). Most of the increase will be in the population aged 45+, and not until after 2020 will the comparative increase in the 65+ age group be greater than that for the 45-64 age group. Disability attributed to arthritis in the population aged 15+ is projected to increase from a prevalence of 2.3% (595,000) in 1991 to 3.3% (1.13 million) in 2031. CONCLUSION: There are large projected increases in both the prevalence and numbers of people with arthritis and arthritis related disability that, at least in the next 20 years, will be split between the older half of the working population and those aged 65 and older.     

Detection of neutralizing antibodies against human T-cell leukemia virus type 1 using a cell-free infection system and polymerase chain reaction

We have developed a cell-free infection system to titrate neutralizing antibodies against human T-cell leukemia virus type 1 (HTLV-1) using the polymerase chain reaction (PCR). S+L-CCC (8C) feline kidney or U-251 MG human glioma cells were infected with a cell-free culture supernatant derived from HTLV-1-infected c77 feline cells. DNA was extracted from 8C or U-251 MG cells after incubation for 24 hr and amplified by PCR. The c77 cell supernatant gave discrete bands, whereas those of HTLV-1-positive T cells did not. When the inocula were treated with HTLV-1 antibody-positive human sera or the monoclonal or polyclonal antibody against the peptide 190-199 of HTLV-1 envelope protein gp46, the subsequent formation of HTLV-1 proviral DNA was inhibited. We determined the titers of neutralizing antibodies by densitometrically scanning the intensity of the PCR bands. These titers correlated well with those determined by the plaque assay using a pseudotype of vesicular stomatitis virus bearing the envelope antigens of HTLV-1. At high serum concentrations, many seronegative samples markedly inhibited the plating of the HTLV-1 pseudotype whereas they barely affected results obtained by PCR. Thus, the c77-PCR system can detect neutralizing antibodies against HTLV-1 even at low titers.     

Human papillomavirus detection in high-grade squamous intraepithelial lesions. Comparison of hybrid capture assay with a polymerase chain reaction system

The validity of human papillomavirus (HPV) detection using the hybrid capture assay (HCA) was compared with the polymerase chain reaction (PCR) in 38 patients with high-grade squamous intraepithelial lesions (HSILs). HCA and PCR showed 84% agreement for HPV detection. HCA missed a significant higher proportion of HSIL compared with PCR (21% vs. 5%; P = .04). Thus, the sensitivity of HCA should be increased before this test can be recommended for HSIL.     

Detection with the polymerase chain reaction of human papillomavirus DNA in condylomata acuminata treated with CO2 laser and microwave

BACKGROUND: The recurrence rates of condyloma acuminata are high. The reasons for the relatively high relapse rates with different treatments are unknown. METHODS: Twelve specimens of condylomata acuminata of the vulva were excised from 12 patients and divided into three parts. One part was untreated, the second and the third parts were treated with CO2 laser and microwave, respectively. DNA was then extracted from tissue by proteolytic digestion and amplified by the polymerase chain reaction. Dot blots were performed with the use of radiolabeled consensus and human papilloma virus (HPV) type-specific probes. RESULTS: HPV DNA was amplified in 100% of untreated specimens (6-HPV 6; 6-HPV 11), and in 83.3% and 50% of specimens treated with CO2 laser and microwave, respectively. There was a significant difference in detection between untreated and microwave-treated specimens (chi 2 = 4.18, P < 0.05). CONCLUSION: Microwave damages HPV DNA more effectively than CO2 laser.     

Demonstration of Epstein-Barr viral DNA in paraffin-embedded tissues of Burkitt's lymphoma from Argentina using the polymerase chain reaction and in situ hybridization

Burkitt's lymphoma (BL) is the most frequent non-Hodgkin's lymphoma in children in Argentina. Although epidemiologic studies have linked Epstein-Barr virus (EBV) to more than 90% of African BL cases but to only 10-20% of American and French cases, increased EBV-specific antibody titers were demonstrated in 73% of Argentinian patients with BL. To characterize the relationship between EBV and BL in Argentina, we analyzed paraffin-embedded tissues from 16 cases of BL for the presence of EBV DNA using the polymerase chain reaction (PCR) and in situ hybridization (ISH). PCR analysis showed that only 4 of 16 specimens contained the EBV BamW fragment, and these specimens were all from cases diagnosed in 1984. Results of ISH performed with a specific biotinylated DNA probe against the NotI/PstI fragment of EBV correlated with the PCR findings. EBV sequences were detected with ISH in 70-90% of the tumor cells from the 4 positive cases. These data may suggest an epidemic outbreak of EBV-related BL in 1984 superimposed on sporadic cases of BL, for which EBV may not have been an essential factor. This study also demonstrates the value of using molecular techniques on archival tissue to track epidemiologic trends.     

Analysis of prostate tissue DNA for the presence of human papillomavirus by polymerase chain reaction, cloning, and automated sequencing

BACKGROUND AND OBJECTIVE: As photorefractive keratectomy (PRK) becomes more widely used, the incidence of repeated PRK increases. The present study was conducted to evaluate results of repeated PRK in view of the meager data on this topic. PATIENTS AND METHODS: In this retrospective study, the authors reviewed the records of 1028 eyes that had undergone PRK, and analyzed the results of 66 eyes that required a second PRK for undercorrection according to baseline refraction. RESULTS: A second PRK was performed in 6.3%, 13.7%, and 10.1% of low, moderate, and high myopes, respectively. The mean refraction 1 year after repeated PRK was similar in both myopic groups: less than -1.00 D. Of the low myopes, 87.50% had residual refraction within 1 D after 1 year. Of the moderate myopes, 88.23% had residual refraction within 1 D after 1 year. All of the low myopes achieved uncorrected visual acuity (VA) of 20/25 or better 1 year after repeated PRK, compared with 58.82% of the moderate myopes. Loss of best-corrected VA never exceeded two lines. CONCLUSION: The overall results of PRK appear to be satisfactory.