Biomedical subjects

Ingo Nindl

Publications and source records attributed to Ingo Nindl.

12 recordsLinked to original sources

PubMed2006

Identification of differentially expressed genes in cutaneous squamous cell carcinoma by microarray expression profiling.

BACKGROUND: Carcinogenesis is a multi-step process indicated by several genes up- or down-regulated during tumor progression. This study examined and identified differentially expressed genes in cutaneous squamous cell carcinoma (SCC). RESULTS: Three different biopsies of 5 immunosuppressed organ-transplanted recipients each normal skin (all were pooled), actinic keratosis (AK) (two were pooled), and invasive SCC and additionally 5 normal skin tissues from immunocompetent patients were analyzed. Thus, total RNA of 15 specimens were used for hybridization with Affymetrix HG-U133A microarray technology containing 22,283 genes. Data analyses were performed by prediction analysis of microarrays using nearest shrunken centroids with the threshold 3.5 and ANOVA analysis was independently performed in order to identify differentially expressed genes (p < 0.05). Verification of 13 up- or down-regulated genes was performed by quantitative real-time reverse transcription (RT)-PCR and genes were additionally confirmed by sequencing. Broad coherent patterns in normal skin vs. AK and SCC were observed for 118 genes. CONCLUSION: The majority of identified differentially expressed genes in cutaneous SCC were previously not described.

Adolescent↗

PubMed2006

Phylogenetic analysis of beta-papillomaviruses as inferred from nucleotide and amino acid sequence data.

Human papillomaviruses (HPV) of the beta-group seem to be involved in the pathogenesis of non-melanoma skin cancer. Papillomaviruses are host specific and are considered closely co-evolving with their hosts. Evolutionary incongruence between early genes and late genes has been reported among oncogenic genital alpha-papillomaviruses and considerably challenge phylogenetic reconstructions. We investigated the relationships of 29 beta-HPV (25 types plus four putative new types, subtypes, or variants) as inferred from codon aligned and amino acid sequence data of the genes E1, E2, E6, E7, L1, and L2 using likelihood, distance, and parsimony approaches. An analysis of a L1 fragment included additional nucleotide and amino acid sequences from seven non-human beta-papillomaviruses. Early genes and late genes evolution did not conflict significantly in beta-papillomaviruses based on partition homogeneity tests (p > or = 0.001). As inferred from the complete genome analyses, beta-papillomaviruses were monophyletic and segregated into four highly supported monophyletic assemblages corresponding to the species 1, 2, 3, and fused 4/5. They basically split into the species 1 and the remainder of beta-papillomaviruses, whose species 3, 4, and 5 constituted the sistergroup of species 2. beta-Papillomaviruses have been isolated from humans, apes, and monkeys, and phylogenetic analyses of the L1 fragment showed non-human papillomaviruses highly polyphyletic nesting within the HPV species. Thus, host and virus phylogenies were not congruent in beta-papillomaviruses, and multiple invasions across species borders may contribute (additionally to host-linked evolution) to their diversification.

Amino Acid Sequence↗

PubMed2006

Identification of dysregulated genes in cutaneous squamous cell carcinoma.

Carcinogenesis is a multi-step process resulting from the accumulation of genetic mutations and subsequently leading to dysregulated genes, but the number and identity of differentially expressed genes in cutaneous squamous cell carcinoma (SCC) is unknown at present. In order to identify dysregulated genes, we examined the relative mRNA expression present in cutaneous SCC and its precursor lesion actinic keratosis (AK) by comparison to normal skin. Snap frozen biopsies from 20 specimens of normal skin, 10 AK, and 10 cutaneous SCC were examined. Total-RNA was extracted, reversely transcribed, and 14 genes were investigated using gene-specific intron-flanking primers and quantitative real-time reverse transcription PCR. Specificity was confirmed by sequencing of the PCR amplicons. Ten of 14 genes were significantly dysregulated in AK and/or cutaneous SCC by comparison to normal skin. The genes CNN2, COX4I1, COX5B, COX7C, CRLF3, CTSC, NDRG1, and LMNA showed increased expression in skin cancer (p < 0.02), while RPL15 and LGTN were down-regulated (p < 0.03). The genes differentially expressed during skin carcinogenesis may prove useful in order to understand the origin and progression of cutaneous SCC and for diagnostic approaches.

Aged↗

PubMed2005

Human papillomavirus-DNA loads in actinic keratoses exceed those in non-melanoma skin cancers.

Recent studies suggest a role of cutaneous human papillomaviruses (HPV) in non-melanoma skin cancer (NMSC) development. In this study viral DNA loads of six frequent HPV types were determined by quantitative, type-specific real-time-PCR (Q-PCR) in actinic keratoses (AK, n=26), NMSC (n=31), perilesional tissue (n=22), and metastases of squamous cell carcinomas (SCC) (n=8) which were previously shown to be positive for HPV5, 8, 15, 20, 24, or 36. HPV-DNA loads in AK, (partially microdissected) NMSC, and perilesional skin ranged between one HPV-DNA copy per 0.02 and 14,200 cell equivalents (median: 1 HPV-DNA copy per 344 cell equivalents; n=48). In 32 of the 79 HPV-positive skin biopsies and in seven of the eight metastases viral loads were even below the detection limit of Q-PCR. Low viral loads in NMSC were confirmed by in situ-hybridization showing only a few HPV-DNA-positive nuclei per section. Viral loads in SCC, basal cell carcinomas, and perilesional tissue were similar. But, viral loads found in AK were significantly higher than in SCC (p=0.035). Our data suggest that persistence of HPV is not necessary for the maintenance of the malignant phenotype of individual NMSC cells. Although a passenger state cannot be excluded, the data are compatible with a carcinogenic role of HPV in early steps of tumor development.

Adult↗

PubMed2005

Development of a general-primer-PCR-reverse-line-blotting system for detection of beta and gamma cutaneous human papillomaviruses.

The beta and gamma genera of papillomaviruses consist of epidermodysplasia verruciformis-related human papillomaviruses (HPVs) and phylogenetically related cutaneous HPVs. Here, we have developed a consensus primer PCR assay and reverse line blot typing system coupled thereto (referred to as beta and gamma cutaneous HPV PCR [BGC-PCR]) for detection and typing of 24 beta and gamma HPVs (HPV types 4, 5, 8, 9, 12, 14, 15, 17, 19, 20, 21, 22, 23, 24, 25, 36, 37, 38, 47, 48, 49, 50, 60, and 65). Because the HPV-specific PCR products are only 72 bp in size, the system is suitable for formalin-fixed, paraffin-embedded specimens and other samples in which the DNA is of suboptimal quality. This system was able to detect and type as little as 100 ag to 1 fg HPV DNA per reaction (depending on the HPV type) in a background of 100 ng human DNA without any cross-reactivity between the tested types. Beta and gamma HPVs were detected in DNA extracted from plucked eyebrow hairs of 31 of 34 renal transplant recipients. In addition, formalin-fixed, paraffin-embedded specimens from nonmelanoma skin tumors of renal transplant recipients (n = 25) and immunocompetent individuals (n = 15) scored BGC-PCR positive in 21 and 6 cases, respectively, with HPV type 5 (HPV5) and HPV8 being the predominant types. The data indicate that this method can be a valuable, user-friendly tool for the detection and typing of cutaneous HPV in clinical specimens and may have implications for future monitoring of vaccines or alternative treatment modalities for diseases caused by these cutaneous HPVs.

Base Sequence↗

PubMed2004

High sensitivity and reproducibility of immunohistochemistry with microagitation.

We examined the practicability, reproducibility and analytical sensitivity of classical immunohistochemistry (IHC) and IHC with microagitation. Two monoclonal antibodies, Ki-67 (proliferation marker) and p53 (tumor suppressor marker), were used. Consecutive paraffin sections of biopsies of suspicious lesions of patients with non-melanoma skin cancer were used in the study. Reproducibility was examined using specimens from four patients in three independent experiments with antibodies against Ki-67 and p53. Analytical sensitivity of the two methods was determined using serial dilutions in two independent experiments. IHC with microagitation could be carried out without destroying the tissue. The new technique was consistent and reproducible, and no background staining was observed. The primary antibodies Ki-67 and p53 could be used at higher dilutions (four to ten times) with microagitation compared with classical IHC. Microagitation can be used for immunohistochemistry; it was reproducible, highly sensitive, and antibodies could be used at higher dilutions. Further analyses with other antibodies using this technique are warranted.

Biopsy↗

PubMed2004

Human papillomaviruses in transplant-associated skin cancers.

BACKGROUND: Human papillomavirus (HPV) infection has been suggested to be involved in the development of nonmelanoma skin cancer, the most common malignancy after solid-organ transplantation. OBJECTIVE: The objective of this study was to analyze the prevalence of different HPV types in squamous cell carcinomas (SCC) and basal cell carcinomas (BCC) of transplant recipients and nonimmunosuppressed patients. METHODS: To include the complete spectrum of HPV types in skin lesions, a comprehensive polymerase chain reaction assay with five different primer combinations was used. RESULTS: For SCC, HPV DNA was detected more frequently in tumors of transplant recipients (12/16, 75%) than of nonimmunosuppressed patients (7/19, 37%). In contrast, the HPV detection rate was similar in BCC specimens (4/8 or 50% in transplanted patients; 27/56 or 48% in nonimmunosupressed patients). Overall, 22 different HPV types were identified. HPV types 5 and 8 were detected predominately in SCC from transplant recipients. The amount of viral DNA was slightly higher in SCC of transplanted than in nonimmunosuppressed patients, but much lower than in both cutaneous and genital warts. CONCLUSIONS: Cutaneous infections with HPV5 and HPV8 may represent an increased risk for SCC development in transplant recipients. The mechanisms by which these viruses may contribute to skin cancer development still remain unclear.

Adult↗

PubMed2004

Human papillomavirus and overexpression of P16INK4a in nonmelanoma skin cancer.

BACKGROUND: P16INK4a overexpression has been identified as a specific biomarker in high-risk human papillomavirus (HPV)-infected cervical (pre)cancer lesions. OBJECTIVE: To evaluate the overexpression of this cyclin-dependent kinase inhibitor in skin tumors depending on HPV infections, we analyzed normal skin, benign skin disease, and skin cancer specimens. METHODS: Biopsies of 23 patients with normal histology (3), psoriasis (2), verrucae vulgaris (2), actinic keratoses (5), squamous cell carcinoma (SCC) in situ (3), Bowen's carcinoma (1), and SCC (7) were analyzed. Specimens of 23 patients were immunostained using the monoclonal antibody E6H4 specific for p16INK4a. HPV status was assessed by a polymerase chain reaction (PCR) system to detect all currently known HPV types. MY (MY09/MY11 and MYN9/MYN10)-, CP (CP65/CP70 and CP66/CP69)-nested PCR, and three single PCR methods CN1, CN3, and CN4 were used in a first step, and HPV typing was performed by restriction fragment length polymorphism analysis. Only beta-globin-positive patients were included in this study. RESULTS: HPV DNA was detected in all actinic keratoses, SCC in situ, Bowen's carcinoma, and SCC, in 50% (one of two) of verrucae vulgaris, in 66% (two of three) of normal skin, and in none of two psoriasis. P16INK4a expression was not detected in normal skin, psoriasis, and verrucae vulgares. Overexpression of p16INK4a was detected in a subset of dysplastic cells (10% to 80%) of all skin (pre)cancer lesions such as actinic keratoses, SCC in situ, Bowen's carcinoma, and SCC infected with HPV independent of sun exposure. CONCLUSION: P16INK4a appears to be overexpressed in a portion of dysplastic cells from actinic keratoses and SCC. Further studies to examine the association of HPV infection and the overexpression of p16INK4a are warranted.

Aged↗

PubMed2003

Assessment of TGF-beta1-mediated growth inhibition of HPV-16- and HPV-18-transfected foreskin keratinocytes during and following immortalization.

The responsiveness to transforming growth factor-beta1 (TGF-beta1) of two human keratinocyte cell lineages (FK16A and FK18B) generated after transfection with HPV-16 and HPV-18, respectively, was investigated. Both cell lineages revealed loss of heterozygosity (LOH) at 18q and/or 3p associated with the acquisition of the immortal phenotype. These loci harbour genes (TGF-beta receptor II gene at 3p, and Smad2 and Smad4 genes at 18q) encoding products involved in the TGF-beta1 signalling pathway. Mortal and early immortal stages of both cell lineages displayed growth reduction upon exposure to TGF-beta1 concentrations in the range 100 pg/ml to 1 ng/ml. However, the late immortal stages were resistant to TGF-beta1 at concentrations up to 10 ng/ml. TGF-beta1 receptors type I and II were expressed at all stages in both cell lineages. Moreover, mRNA levels of Smad2 and Smad4 genes were nearly constant throughout. TGF-beta1 expression and secretion, which were demonstrated in all analysed stages, may provide selective conditions underlying unresponsiveness to TGF-beta1 upon prolonged monolayer culturing. Thus, LOH at 3p and/or 18q seen during HPV-mediated immortalization of human keratinocytes was not associated with resistance to TGF-beta1-mediated growth inhibition or a marked reduction in TGF- beta1 receptors and mRNA levels of Smad2 or Smad4. Therefore, alternative events are likely to underlie unresponsiveness to TGF- beta1 in late-passage FK16A and FK18B cells.

Cell Division↗

PubMed2003

Association of human papillomavirus infections with cutaneous tumors in immunosuppressed patients.

Besides immunosuppression and UV radiation, human papillomavirus (HPV) infection was also suggested to be involved in the development of non-melanoma skin cancer, the most common malignancy after transplantation. In this study we used a comprehensive PCR assay to analyze the prevalence of individual HPV types in different skin lesions from transplant and non-transplant patients. HPV DNA was detected more frequently in squamous cell carcinomas (SCCs) of transplant recipients (75%) than the same lesion was in non-immunosuppressed patients (47%). Similar HPV prevalences were found in cutaneous warts (91% vs 94%), pre-malignant skin tumors (38% vs 36%), and normal skin specimens (17% vs 16%) of both patient populations. Overall, more than 40 different HPV types were identified. HPV types 5 and 8 were found more frequently in SCCs (26%) than in pre-cancerous (5%) or benign lesions (1%). All HPV 5- and HPV 8-positive SCCs were from immunosuppressed patients, indicating that infection with HPV 5 and HPV 8 may present an increased risk of SCC development in these patients.

Base Sequence↗

PubMed2003

A linkage map of an F2 hybrid population of Antirrhinum majus and A. molle.

To increase the utility of Antirrhinum for genetic and evolutionary studies, we constructed a molecular linkage map for an interspecific hybrid A. majus x A. molle. An F(2) population (n = 92) was genotyped at a minimum of 243 individual loci. Although distorted transmission ratios were observed at marker loci throughout the genome, a mapping strategy based on a fixed framework of codominant markers allowed the loci to be placed into eight robust linkage groups consistent with the haploid chromosome number of Antirrhinum. The mapped loci included 164 protein-coding genes and a similar number of unknown sequences mapped as AFLP, RFLP, ISTR, and ISSR markers. Inclusion of sequences from mutant loci allowed provisional alignment of classical and molecular linkage groups. The total map length was 613 cM with an average interval of 2.5 cM, but most of the loci were aggregated into clusters reducing the effective distance between markers. Potential causes of transmission ratio distortion and its effects on map construction were investigated. This first molecular linkage map for Antirrhinum should facilitate further mapping of mutations, major QTL, and other coding sequences in this model genus.

Antirrhinum↗

PubMed2003

Correlation of p16INK4A overexpression with human papillomavirus infection in cervical adenocarcinomas.

SUMMARY: As human papillomavirus (HPV) infection is the main risk factor for squamous cell carcinoma of the cervix and overexpression of p16INK4a occurs when retinoblastoma protein is inactivated by high-risk HPV, the authors studied the association of HPV infection and expression of p16INK4a in cervical adenocarcinomas. Specimens of cervical glandular neoplasias were immunostained with a p16INK4a-specific monoclonal antibody (clone E6H4). Approximately 80% of glandular neoplasms showed overexpression of p16INK4a. Exfoliated cells from 14 adenocarcinomas were further examined by p16INK4a-specific immunocytochemistry, and 12 cases showed overexpression of p16INK4a, suggesting that immunostaining for p16INK4a may be a useful diagnostic tool for cervical adenocarcinomas. The authors further examined HPV DNA in cervical adenocarcinomas with the polymerase chain reaction method. Overexpression of p16INK4a was positive in 94% of cases in which HPV16 or 18DNA was positive, a finding suggesting that HPV16 or 18 may play an important role in cervical adenocarcinomas. Overexpression of p16INK4a may be an indicator of pathogenic activity of high-risk HPVs.

Adenocarcinoma↗