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Biomedical subjects

Y Chardonnet

Publications and source records attributed to Y Chardonnet.

At least 19 recordsLinked to original sources
PubMed1992

Epidermotropism of T cells correlates with intercellular adhesion molecule (ICAM1) expression in human papillomavirus (HPV)-induced lesions.

Adhesion molecules play an important role in inflammatory reactions. Among them, ICAM1, a ligand for the lymphocyte function-associated antigen (FLA1) of leucocytes, may be expressed by antigen-presenting cells and keratinocytes in various inflammatory disorders. As cell-mediated immune responses play a great role in HPV infections, we investigated the expression of ICAM1 and correlated it with the presence of LFA1-positive cells by immunohistochemistry on serial frozen sections of a series of non-regressing cutaneous and mucosal HPV-induced lesions. ICAM1 expression by keratinocytes was observed only in intensely infiltrated lesions of condylomas and laryngeal papillomas. Its induction was usually correlated with the presence of LFA1-positive cells (mainly CD8-positive cells) which were in close apposition to ICAM1-positive proliferative epithelial cells expressing also, in some cases, HLA-DR antigen. ICAM1 was not correlated with the presence of HPV DNA or viral antigen. In moderately infiltrated lesions, keratinocytes did not express ICAM1, and LFA1-positive cells were not observed in the epidermis. In all lesions, ICAM1 was more intense on endothelial cells than in normal skin; infiltrating cells (lymphocytes and dendritic cells) may also express this antigen but intraepithelial Langerhans cells were devoid of any labelling. These studies provide further evidence that T-lymphocyte mechanisms are important in the host response to HPV-induced lesions. ICAM1 expression correlates with a lesional infiltrate but not with HPV infection and probably results in a more efficient initiation of the immune reaction.

CD4 Antigens↗
PubMed1992

Detection of multiple types of human papillomavirus in a giant condyloma from a grafted patient. Analysis by immunohistochemistry, in situ hybridisation, Southern blot and polymerase chain reaction.

Immunosuppressed patients such as transplant recipients are known to develop multiple lesions suggestive of human papillomavirus (HPV) infection. A giant anal condyloma was obtained from a transplant patient; several fragments taken from different areas were examined for the presence of HPV DNA using in situ hybridisation, polymerase chain reaction (PCR) and Southern blot. Typical koilocytes were seen in routinely stained tissue sections, suggesting an HPV infection; furthermore, group specific HPV antigen was detected in one of four frozen fragments. Different results were obtained by in situ hybridisation according to the fragment tested. HPV types 6/11 were detected in each of the five fragments, frozen or fixed in Bouin's or formalin solutions. However, the number of HPV DNA positive cells and the intensity of the reaction greatly varied with the specimen. HPV 16 and 18 probes also reacted positively with the sample fixed in formalin; a stronger signal was observed with HPV 18 in one large focus than with HPV 16. HPV type 5 was detected in a few isolated cells of two frozen fragments. With the Southern blot technique, the profile of an HPV 6/11 was seen only in one of two frozen fragments; in this case, the bands were intense. A slight positive reaction was also obtained in one frozen fragment with HPV 16 probe. Four frozen fragments were analyzed with PCR: HPV 6/11 was detected in each fragment; HPV 18 was detected in the four samples but with different intensities; HPV types 5 and 16 did not show any positive signal. In conclusion, the lesion is an example of infection with several HPV types, demonstrated by three different techniques. This suggests the need for careful dermatological or colposcopic follow-up of transplant recipients, in order to prevent possible malignant transformation of anogenital lesions.

Adolescent↗
PubMed1992

c-myc and c-Ha-ras cellular oncogenes and human papillomaviruses in benign and malignant cutaneous lesions.

To analyze the role of human papillomavirus (HPV) infection and c-myc and c-Ha-ras oncogene activation in cutaneous and mucosal lesions, serial frozen sections of 47 lesions from grafted recipients and 10 biopsies from non-immunosuppressed patients were examined. HeLa, CaSki, MCF7, Colo 320 and 3T3 cells, which contain various copy numbers of HPV DNA and/or c-myc gene, were used as controls. HPV, myc and ras oncogene DNAs were not detected in normal epithelia by in situ hybridization with biotinylated DNA probes. The amplification of ras oncogene was detected in 20/57 lesions. The amplification of myc oncogene was found in 14/57 lesions, 13 of which showed both myc and ras gene amplification. c-myc and/or c-Ha-ras DNA was more frequently amplified in cutaneous squamous cell carcinomas (8/14 cases) and anogenital papillomas (4/6 cases), than in common and plantar warts (3/14 cases) or actinic keratoses (2/10 cases). HPV DNA was detected in 26/57 biopsies. Oncogene amplification was codetected with HPV DNA in 10/26 lesions, each of them containing at least one potentially oncogenic HPV type (5,16 and/or 18). The amplification was also found in 11/31 cases in the absence of HPV DNA. No significant difference was observed in the detection of HPV or oncogene DNA between the lesions of transplant recipients and those of the non-immunosuppressed population. Viral antigen was detected in 17/55 lesions by indirect immunofluorescence; 5 of the positive biopsies showed ras oncogene amplification. Myc and ras p21 oncoproteins were respectively localized in the nuclei and on the membrane of epithelial cells by indirect immunofluorescence. A good correlation was observed between the amplification of oncogenes and the expression of oncoproteins. Our results favor the hypothesis of a cooperation between HPV infection and myc and ras oncogene activation in skin carcinogenesis.

Blotting, Southern↗
PubMed1992

Human papillomavirus DNA in cervix. In-situ hybridization with biotinylated probes on Bouin's fixed paraffin embedded specimens.

We examined retrospectively a series of 65 Bouin's fixed, paraffin-embedded tissue specimens from 8 condylomatous lesions, 16 condylomas associated with cervical intraepithelial neoplasia (CIN), and 12 neoplasia without condylomatous signs, for histological characteristics, the detection of viral structural antigen, the presence and typing of HPV DNA by molecular in situ hybridization with biotinylated probes types 6, 11, 16 and 18 under stringent conditions (Tm - 12 degrees C). HPV DNA was present in 34/65 (52%) specimens. Detection of viral structural antigen was positive in only 14% (3/22) specimens. HPV DNA were identified in 9/9 (100%) condylomatous lesions (with HPV type 6, 11, 18). Three condylomas were coinfected with both HPV type 6 or 11 and type 18; viral antigen was found in two specimens. HPV DNA were detected in 18/31 (58%) low grade and advanced CIN associated with condylomatous changes (type 6 = 5 specimens, type 11 = 3 specimens, type 16 = 4 specimens, type 18 = 6 specimens). Four of these cases were coinfected with both HPV type 6/11 and HPV type 16/18. Viral antigen was negative in all specimens. HPV DNA were detected in 7/25 (28%) advanced intra-cervical neoplasia (CIN III) without anatomopathological condylomatous changes (type 6 = 1 specimen, type 16 = 3 specimens, type 18 = 3 specimens). One of these specimens contained both HPV types 6 and 18. Viral antigen was found in one case. Our data confirm the association of HPV types 6 and 11 with condyloma and low grade neoplasia; HPV types 16 and 18 were associated with advanced cervical neoplasia.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetates↗
PubMed1992

Influence of fixation on human papillomavirus DNA detection in frozen and embedded paraffin lesions by in situ hybridization with biotinylated probes.

Series of frozen or paraffin-embedded tissues from various body sites, taken from non-immunosuppressed or immunosuppressed patients with persistent papilloma lesions were examined for the presence of group specific antigen from human papillomavirus (HPV) by indirect immunofluorescence or HPV DNA by in situ hybridization with biotinylated probes. We have shown that it is possible to detect HPV DNA after fixation of tissues in neutral formalin, Bouin's or Baker's solution. However, the sensitivity was reduced as compared to frozen tissues. The HPV DNA was detected in nuclei of heavily infected epithelial cells such as plantar or hand warts or in dispersed cells containing high copy numbers of HPV DNA from lesions such as squamous cell carcinomas or keratoacanthomas. In premalignant or malignant lesions of both immunosuppressed or non-immunosuppressed patients, HPV DNA was rarely detected after fixation. HPV types commonly described for skin and genital samples were identified in non-immunosuppressed patients, whereas in transplant recipients oncogenic HPV type 16 was identified in benign warts as well as in premalignant or malignant lesions. Positive reactions with several HPV types were more frequent in lesions from grafted patients than from the normal population. Virus antigen was detectable more frequently in frozen sections than in fixed tissues. Our findings indicate that in situ hybridization is an appropriate and rapid technique to study the presence of HPV infection. However, numerous controls are needed to avoid misinterpretations.

Antigens, Viral↗
PubMed1992

Langerhans cells, inflammation markers and human papillomavirus infections in benign and malignant epithelial tumors from transplant recipients.

Organ transplant recipients frequently develop warts which progress toward premalignant or malignant lesions after a rather long grafting period. The local immune responses of such lesions (warts, condyloma acuminata, actinic keratoses, Bowen, basal and squamous cell carcinomas) was studied in 32 frozen skin specimens taken from 15 male transplant recipients and compared to similar lesions from the normal population. We studied the expression of T cell subsets, Langerhans cell phenotype, HLA class 1 (beta 2-microglobulin), HLA class 2 (DR antigen), and intercellular adhesion molecule 1 (ICAM 1). The presence of HPV infection was also considered, using in situ hybridization with biotinylated probes in order to examine the correlation with immunological markers. In the dermis, the lesions from grafted patients showed a moderate to intense inflammatory reaction of HLA-DR-positive cells. Most of these cells were CD4+ and CD8+ without any predominance of a single T cell subset. In the epidermis, most lesions were characterized by a reduced number of CD1-positive cells; this was concomitant with a decrease or a loss of beta 2-microglobulin expression by epithelial cells. HLA-DR antigen was not expressed by keratinocytes or tumoral cells in any specimen; ICAM 1 antigen was observed in a few cases. The expression of these markers was similarly modified with or without the presence of HPV DNA. Conversely, most lesions from non-immunocompromised patients, except warts, showed intense inflammatory reactions, with a predominance of CD4-positive cells and large foci of ICAM 1-positive cells. Expression of activation markers by keratinocytes occurred mainly in condylomas and squamous cell carcinomas. In the normal population, HPV infection was only detected in papilloma lesions. These data indicate, in lesions from grafted patients, a lack of effective immune response with partial inhibition of activation markers expressed by keratinocytes. It is conceivable that immunosuppressive treatment with solar exposure may also be responsible for the local immune deficiency and thus for the conversion of benign warts toward malignant lesions in grafted patients.

Adult↗
PubMed1992

Human papillomavirus detection in cervical cells by in situ hybridization with biotinylated probes.

We have investigated the applicability of human papillomavirus (HPV) DNA detection by in situ hybridization with biotinylated probes in epithelial cells obtained from the cervix using a cotton tip swab. We describe a simple procedure for obtaining homogeneous cell samples and good preservation of cellular structure. This is achieved by pretreatment of cells with L-cysteine before hybridization. Separate denaturation of cellular DNA and probe DNA is also necessary for satisfactory results. Both benign HPV DNA 6/11 and potentially oncogenic HPV DNA 16/18 could be identified in our series. In situ hybridization on cervical scrapes is a rapid, simple and very specific method for detecting patients infected with oncogenic HPV types.

Adult↗
PubMed1992

Evaluation of human papillomavirus type 5 on frozen sections of multiple lesions from transplant recipients with in situ hybridization and non-isotopic probes.

Transplant recipients are at high risk to develop multiple cutaneous lesions after grafting. The frequency of the potentially oncogenic human papillomavirus (HPV) type 5 DNA was evaluated in cutaneous lesions taken from sun-exposed areas in transplant recipients (92 lesions and 5 samples from normal skin) and compared with a nontransplanted population (22 lesions and 7 samples from normal skin) using in situ hybridization and biotinylated probes to HPV types 1, 2, 5, 16 and 18. HPV type 5 DNA was identified in 8/92 cutaneous lesions of transplanted recipients: 3 warts, 1 case of seborrheic keratosis, 2 actinic keratoses and 2 keratoacanthomas. HPV type 5 DNA was not detected in 27 malignant tumors (8 basal cell carcinomas and 19 squamous cell carcinomas) from transplant recipients. HPV DNA type 5 was detected in only 1 case of squamous cell carcinoma from the general population. The presence of HPV DNA 5 was confirmed with Southern blotting in 2 out of 6 cases from transplant recipients. The reaction was negative with the squamous cell carcinoma from nontransplant recipients. These data indicate that the presence of HPV DNA type 5 is not very frequent; it can be detected with in situ hybridization and nonisotopic probe, which is easier to handle than Southern blot.

Biopsy↗
PubMed1992

[Malpighian epithelia and papillomavirus infections].

Human papillomaviruses (HPV) are a large group of DNA viruses, with over 60 types identified to date, which can cause the development of benign tumors in the skin and mucosal squamous epithelia. Most of these tumors regress spontaneously but some, especially in the mucosal membranes, become malignant. HPV types with a high risk for inducing malignancies (e.g. 16 and 18) are the subject of increasing interest. HPVs are both host-specific and tissue-specific: some types preferentially infect specific epithelia, giving rise to lesions with distinct topographic characteristics. HPVs are difficult to study because they do not replicate in available in vitro models. In vivo, HPVs replicate well in epithelial cells undergoing terminal differentiation, e.g. in keratinized cells. Some 40 different types have been reported in epidermal keratinocytes, the most common being types 1 and 2 which produce large amounts of viral antigens and viral particles. In contrast, HPVs replicate poorly in the weakly keratinized squamous epithelia which line the digestive, respiratory, and genital tracts. Junctional epithelia, e.g. on the uterine cervix, are especially prone to HPV infection. The most prevalent HPV types in benign genital lesions are types 6 and 11, whose characteristic features include extrachromosomal DNA and production of only small amounts of viral antigens. The profound nuclear and cytoplasmic changes induced by HPVs lead to the formation of koïlocytes which are found mainly in the granular layer of epithelia and have been especially well described in the uterine cervix and vagina. HPV epithelial tumors are squamous cell carcinomas that often harbor HPV types 16 and 18; this is especially true of cervical intraepithelial neoplasias.(ABSTRACT TRUNCATED AT 250 WORDS)

Anus Diseases↗
PubMed1992

[Human papillomaviruses and cellular oncogenes (c-myc, c-Ha-ras) in cutaneous and mucosal lesions in transplantation recipients].

Transplant recipients develop numerous benign and malignant cutaneous and mucosal lesions in which histological signs of human papillomavirus (HPV) infection are observed. To investigate the role of HPV and c-myc and c-Ha-ras cellular oncogenes' activation in transplanted patients lesions, we used in situ hybridization with biotinylated probes and Southern blot to detect HPV and oncogenes DNA. We analyzed 36 lesions from grafted patients: 11 common warts, 10 actinic keratoses, 13 squamous cell carcinomas and 2 anogenital papillomas. With in situ hybridization, HPV DNA was detected in 14/36 lesions, 10 of which contained several HPV types. Benign and potentially oncogenic HPV types were detected in warts as well as in squamous cell carcinomas. The Southern blot technique confirmed the distribution of HPV types. Group specific viral antigen was detected in 12 lesions, mainly warts. C-Ha-ras oncogene was amplified in 13 lesions and c-myc oncogene in 10 lesions, 9 of which showed both oncogene amplification. The results obtained with in situ hybridization for c-myc gene amplification were confirmed with the Southern blot technique in 11/14 cases. Ras and/or myc amplification was more frequent in squamous cell carcinomas and anogenital papillomas than in warts and actinic keratoses. The amplification was not always linked to the presence of HPV DNA; however, it was more frequent in lesions infected by potentially oncogenic HPV types than in lesions containing only benign HPV types. Myc and p21 oncoproteins were respectively localized in the nucleus and on the membrane of epithelial cells by immunofluorescence. Most lesions showed a good concordance between the detection of oncogene DNA and proteins. Our results suggest than c-Ha-ras and c-myc cellular oncogenes' activation and HPV infection could play a role in the cancerization of cutaneous lesions from transplant recipients.

Genes, myc↗
PubMed1992

[Malpighian epithelia and papillomavirus infections].

Human papillomaviruses (HPV) are a large group of DNA viruses, with over 60 types identified to date, which can cause the development of benign tumors in the skin and mucosal squamous epithelia. Most of these tumors regress spontaneously but some, especially in the mucosal membranes, become malignant. HPV types with a high risk for inducing malignancies (e.g. 16 and 18) are the subject of increasing interest. HPVs are both host-specific and tissue-specific: some types preferentially infect specific epithelia, giving rise to lesions with distinct topographic characteristics. HPVs are difficult to study because they do not replicate in available in vitro models. In vivo, HPVs replicate well in epithelial cells undergoing terminal differentiation, e.g. in keratinized cells. Some 40 different types have been reported in epidermal keratinocytes, the most common being types 1 and 2 which produce large amounts of viral antigens and viral particles. In contrast, HPVs replicate poorly in the weakly keratinized squamous epithelia which line the digestive, respiratory, and genital tracts. Junctional epithelia, e.g. on the uterine cervix, are especially prone to HPV infection. The most prevalent HPV types in benign genital lesions are types 6 and 11, whose characteristic features include extrachromosomal DNA and production of only small amounts of viral antigens. The profound nuclear and cytoplasmic changes induced by HPVs lead to the formation of koïlocytes which are found mainly in the granular layer of epithelia and have been especially well described in the uterine cervix and vagina. HPV epithelial tumors are squamous cell carcinomas that often harbor HPV types 16 and 18; this is especially true of cervical intraepithelial neoplasias. These tumors contain the viral DNA, which may or may not be integrated into the cellular DNA, whereas viral antigens are lacking. The high incidence and broad spectrum of HPV types found in patients with acquired immunodeficiencies (e.g. under immunosuppressive therapy) suggest a key role for cellular immunity in the development of HPV-induced lesions. A number of cofactors, including UV radiations and smoking, as well as oncogene activation and anti-oncogene inactivation, may increase the risk of progression to malignancy.

Digestive System Diseases↗
PubMed1991

Cell fusion of human and mouse cells as a source for new cells retaining human markers. Analysis of DNA content, membrane and cytoplasmic antigen expression.

Flow cytometry and ultrastructural morphometry were used to study some characteristics of cells obtained by fusion with polyethylene-glycol 4000 between mouse fibroblasts 3T3.4E and normal keratinocytes (3T3.4E x NHK) or hand wart human keratinocytes (3T3.4E x HWK), at late passages. The cell cycle and the expression of human beta 2-microglobulin, human EGF-receptors (EGF-r), vimentin were simultaneously studied by flow cytometry. Epithelial CaSki cells, derived from a human uterine carcinoma, expressing high levels of beta 2-microglobulin, EGF-r and vimentin, were used as a positive control. In mouse fibroblasts 3T3.4E only vimentin was expressed whereas in cells derived from fusion, human beta 2-microglobulin, human EGF-r and vimentin were detected. The cell cycle analysis revealed that the peak position of G0/G1 differed with the cells (channel 11 for 3T3.4E cells, 13 for 3T3.4E x HWK and 15 for 3T3.4E x NHK). The area of the cell compartments from each cell type was also different by quantitative ultrastructural morphometry. The hybrid phenotype was maintained in late passages in cells (3T3.4E x NHK) and (3T3.4E x HWK), as shown by the expression of human antigens, differences in DNA contents and nuclear area. Flow cytometry may be a very accurate and precise tool for studying low antigenic expression. The combination of different methods including analysis of DNA content, antigenic expression and ultrastructural morphometry confirmed that 3T3.4E, 3T3.4E x NHK and 3T3.4E x HWK cells are different cell types. These techniques are complementary to cell phenotype analysis.(ABSTRACT TRUNCATED AT 250 WORDS)

3T3 Cells↗
PubMed1991

Human papillomavirus type 1-associated squamous cell carcinoma in a heart transplant recipient.

The occurrence of squamous cell carcinomas in organ transplant recipients with warts represents a good model to study viral carcinogenesis. Most of the cases were reported in renal transplant recipients. We present the case of a heart transplant recipient in whom multiple common warts, preepitheliomatous keratoses, and squamous cell carcinomas developed. The warts began 4 years after the transplantation and the first carcinoma occurred 2 years after the warts, all the lesions being on sun-exposed areas. Histologic signs of human papillomavirus infection were seen in all premalignant and malignant lesions. Furthermore, human papillomavirus type 1 DNA was detected by in situ molecular hybridization within one of the carcinomas. Human papillomaviruses, along with other carcinogenic factors, play an important role in the development of carcinomas, and benign types could be implicated. Further studies are required to evaluate the frequency of cutaneous malignant neoplasms in heart transplant recipients as compared with renal transplant recipients.

Adult↗
PubMed1990

Evaluation of mitochondrial content and activity with nonyl-acridine orange and rhodamine 123: flow cytometric analysis and comparison with quantitative morphometry. Comparative analysis by flow cytometry and quantitative morphometry of mitochondrial content and activity.

Mouse fibroblasts 3T3.4E and two cells lines obtained by fusion (3T3.4E cells x normal human keratinocytes), (3T3 x NHK), and (3T3.4E cells x hand wart keratinocytes), (3T3 x HWK), were compared for mitochondrial activity and content between 5 and 20 days of culture, from the 16th to 20th passage, by using Rh 123 and NAO respectively. In 3T3.4E cells both Rh 123 and NAO fluorescence were similar after 5 and 7 days of culture, indicating no modification of mitochondrial activity and content at that time. However, in cells derived from fusion of 3T3 x NHK or 3T3 x HWK, Rh 123 increased from 5 to 20 days whereas NAO fluorescence was maximal at 7 days of culture and then decreased, indicating that their mitochondrial activity differed from that of 3T3.4E cells. No difference was observed between the 16th and 20th passage. Quantitative morphometry and flow cytometry gave good correlations at 7 days of culture for the cell size, estimated either by the cell area or the cell diameter, and for mitochondria content, evaluated either by the number of mitochondria per cell or NAO fluorescence intensity.

Acridine Orange↗
PubMed1990

Human papillomavirus type 11DNA in papillary squamous cell lung carcinoma.

We report a case of papillary squamous cell carcinoma of the lung developing in relation to a condylomatous papilloma and related to human papillomavirus (HPV) infection. The viral origin of the bronchial papillomatous lesion is strongly suggested by cytological and histological features with marked condylomatous changes. No viral capsid antigen was detected by immunohistochemistry. Transmission electron microscopy failed to reveal intranuclear viral-like particles in the papillary part of the carcinoma, but typical ultrastructural koilocytotic cells with irregular nucleus and coarse chromatin were observed. HPV DNA type 11 was detected by in situ hybridization using biotinylated probes on paraffin-embedded specimens, under stringent conditions (Tm-12 degrees, 50% formamide). Papillary squamous cell carcinoma may result from the malignant conversion of benign squamous papilloma of the bronchus. HPV type 11 may be associated with malignant conversion of benign papilloma of the pulmonary tract, as in the upper respiratory tract. In situ hybridization with biotinylated probes is a relatively simple and appropriate method for retrospective analysis of HPV DNA sequences in surgical specimens.

Bronchial Neoplasms↗