Biomedical subjects

J Mollenhauer

Publications and source records attributed to J Mollenhauer.

At least 19 recordsLinked to original sources

PubMed2006

Molecular characterisation of non-absorptive and absorptive enterocytes in human small intestine.

BACKGROUND AND AIMS: Perturbation of differentiation of the crypt-villus axis of the human small intestine is associated with several intestinal disorders of clinical importance. At present, differentiation of small intestinal enterocytes in the crypt-villus axis is not well characterised. SUBJECTS AND METHODS: Expression profiling of microdissected enterocytes lining the upper part of crypts or the middle of villi was performed using the Affymetrix X3P arrays and several methods for confirmation. RESULTS: A total of 978 differentially expressed sequences representing 778 unique UniGene IDs were found and categorised into four functional groups. In enterocytes lining the upper part of crypts, cell cycle promoting genes and transcription/translation related genes were predominantly expressed, whereas in enterocytes lining the middle of villi, high expression of cell cycle inhibiting genes, metabolism related genes, and vesicle/transport related genes was found. CONCLUSION: Two types of enterocytes were dissected at the molecular level, the non-absorptive enterocyte located in the upper part of crypts and the absorptive enterocyte found in the middle of villi. These data improve our knowledge about the physiology of the crypt-villus architecture in human small intestine and provide new insights into pathophysiological phenomena, such as villus atrophy, which is clinically important.

Adult↗

PubMed2006

Qualitative evaluation of titanium implant integration into bone by diffraction enhanced imaging.

Diffraction enhanced imaging (DEI) uses refraction of x-rays at edges, which allows pronounced visualization of material borders and rejects scattering which often obscures edges and blurs images. Here, the first evidence is presented that, using DEI, a destruction-free evaluation of the quality of integration of metal implants into bone is possible. Experiments were performed in rabbits and sheep with model implants to investigate the option for DEI as a tool in implant research. The results obtained from DEI were compared to conventional histology obtained from the specimens. DE images allow the identification of the quality of ingrowth of bone into the hydroxyapatite layer of the implant. Incomplete integration of the implant with a remaining gap of less than 0.3 mm caused the presence of a highly refractive edge at the implant/bone border. In contrast, implants with bone fully grown onto the surface did not display a refractive signal. Therefore, the refractive signal could be utilized to diagnose implant healing and/or loosening.

Animals↗

PubMed2006

Collagen degradation products modulate matrix metalloproteinase expression in cultured articular chondrocytes.

Destruction of collagen within osteoarthritic cartilage depends in part on collagen-degrading matrix metalloproteases (MMP). Degradative fragments of type II collagen (Col II) occur in normal and in osteoarthritic cartilage, and may contribute to regulation of matrix turnover by interfering with normal cell-matrix communication pathways. Therefore, the effects of different types of collagen fragments on mRNA and protein levels of MMP-2, MMP-3, MMP-9, and MMP-13 in cultured bovine articular knee chondrocytes and explants were examined. Primary chondrocytes and explants were incubated with fragments from whole cartilage collagen matrix (Colf) and from purified type II collagen (Col2f), or with a synthetic 29-mer peptide representing the amino-terminal domain of type II collagen (Ntelo). Gelatin zymography revealed increases of proMMP-2, a shift towards active MMP-2 and increases in proMMP-9, depending on the type of fragment. In situ hybridization of cartilage sections displayed MMP-3 mRNA in virtually all cells. Moderate to strong increases in MMP-2, MMP-3, MMP-9, and MMP-13 mRNA levels were detected by quantitative PCR. The results demonstrate stimulating effects of collagen fragments on both mRNA and/or protein from MMP -2, -3, -9, and -13, and suggest a novel mechanism of MMP induction and activation that includes a particular role for N-telo in controlling catabolic pathways of matrix turnover.

Amino Acid Sequence↗

PubMed2005

Osteointegration of hydroxyapatite-titanium implants coated with nonglycosylated recombinant human bone morphogenetic protein-2 (BMP-2) in aged sheep.

Osteointegration of metal implants into aged organisms can be severely compromised due to reduced healing capacity of bone, lack of precursor cells for new bone formation, or osteoporosis. Here, we report on successful implant healing in a novel model of aged sheep in the presence of nonglycosylated bone morphogenetic protein 2 (BMP-2). Ewes of 8 to 12 years with significant radiologic and histologic signs of osteoporosis and adipocytic bone marrow received a cylindrical hydroxyapatite-titanium implant of 12 x 10 mm. BMP-2 has been produced as a bacterial recombinant fusion protein with maltose-binding protein and in vitro generation of mature BMP-2 by renaturation and proteolytic cleavage. A BMP-2 inhibition ELISA was developed to measure the in vitro release kinetics of bioactive human BMP-2 from immersed solid implant materials by using Escherichia coli expressed and biotinylated recombinant human BMP-2 receptor IA extracellular domain (ALK-3 ECD). The implants were placed laterally below both tibial plateaus, with the left leg implant carrying 380 microg BMP-2. Both implant types became integrated within the following 20 weeks. The control implant only integrated at the cortical bone, and little new bone formation was found within the pre-existing trabecular bone or the marrow cavity. Marrow fat tissue was partially replaced by unspecific connective tissue. In contrast, BMP-2-coated implants initiated significant new bone formation, initially in trabecular arrangements to be replaced by cortical-like bone after 20 weeks. The new bone was oriented towards the cylinder. Highly viable bone marrow appeared and filled the lacunar structures of the new bone. In mechanical tests, the BMP-2-coated implants displayed in average 50% higher stability. This animal model provided first evidence that application of nonglycosylated BMP-2 coated on solid implants may foster bone healing and regeneration even in aged-compromised individuals.

Aging↗

PubMed2005

DMBT1 expression and glycosylation during the adenoma-carcinoma sequence in colorectal cancer.

The gene DMBT1 (deleted in malignant brain tumour-1) has been proposed to play a role in brain and epithelial cancer, but shows unusual features for a classical tumour-suppressor gene. On the one hand, DMBT1 has been linked to mucosal protection, whereas, on the other, it potentially plays a role in epithelial differentiation. Thus its function in a particular tissue is of mechanistic importance for its role in cancer. Because the former function requires secretion to the lumen and the latter function may depend on its presence in the extracellular matrix, we decided to investigate DMBT1 expression, location and its mode of secretion during malignant transformation in colorectal cancer. Using human colorectal PC/AA cell lines and tissue sections from individual patients, we have examined the expression of DMBT1 and its glycosylation in the adenoma-carcinoma sequence leading to the adenocarcinoma phenotype.

Adenocarcinoma↗

PubMed2004

DMBT1 expression is down-regulated in breast cancer.

BACKGROUND: We studied the expression of DMBT1 (deleted in malignant brain tumor 1), a putative tumor suppressor gene, in normal, proliferative, and malignant breast epithelium and its possible relation to cell cycle. METHODS: Sections from 17 benign lesions and 55 carcinomas were immunostained with anti DMBT1 antibody (DMBTh12) and sections from 36 samples, were double-stained also with anti MCM5, one of the 6 pre-replicative complex proteins with cell proliferation-licensing functions. DMBT1 gene expression at mRNA level was assessed by RT-PCR in frozen tissues samples from 39 patients. RESULTS: Normal glands and hyperplastic epithelium in benign lesions displayed a luminal polarized DMBTh12 immunoreactivity. Normal and hyperplastic epithelium adjacent to carcinomas showed a loss of polarization, with immunostaining present in basal and perinuclear cytoplasmic compartments. DMBT1 protein expression was down-regulated in the cancerous lesions compared to the normal and/or hyperplastic epithelium adjacent to carcinomas (3/55 positive carcinomas versus 33/42 positive normal/hyperplastic epithelia; p = 0.0001). In 72% of cases RT-PCR confirmed immunohistochemical results. Most of normal and hyperplastic mammary cells positive with DMBTh12 were also MCM5-positive. CONCLUSIONS: The redistribution and up-regulation of DMBT1 in normal and hyperplastic tissues flanking malignant tumours and its down-regulation in carcinomas suggests a potential role in breast cancer. Moreover, the concomitant expression of DMTB1 and MCM5 suggests its possible association with the cell-cycle regulation.

Agglutinins↗

PubMed2004

Salivary agglutinin/DMBT1SAG expression is up-regulated in the presence of salivary gland tumors.

Salivary agglutinin (SAG) is encoded by the gene Deleted in Malignant Brain Tumors 1 (DMBT1) and represents the salivary variant of DMBT1 (DMBT1(SAG)). While SAG is a bona fide anti-caries factor, DMBT1 was proposed as a candidate tumor-suppressor for brain, digestive tract, and lung cancer. Though DMBT1(SAG) is expressed in the salivary glands, its expression in salivary gland tumors is unknown. Here we analyzed DMBT1(SAG) expression in 20 salivary gland tumors and 14 tumor-flanking tissues by immunohistochemistry. DMBT1(SAG) in salivary gland tumors resembles the changes of expression levels known from DMBT1 in tumors in other cancer types. Particularly, DMBT1(SAG) was up-regulated in 10/14 tumor-flanking tissues, and a strong staining of the luminal content in the tumor and/or the tumor-flanking tissue was observed in 14/20 cases. This suggests that, in addition to its role in caries defense, SAG may serve as a potential tumor indicator and/or tumor suppressor in salivary gland tissue.

Agglutinins↗

PubMed2003

Overwhelming septic infection with a multi-resistant Staphylococcus aureus (MRSA) after total knee replacement.

INTRODUCTION: The incidence of early deep infection after arthroplasty of the knee is very low but could represent a serious future problem. MATERIAL AND METHODS: The authors report on a 71-year-old woman with gonarthritis who was supplied with a total knee endoprosthesis and developed a local infection followed by septic shock. RESULTS: The infection was evidently caused by a multiresistant Staphylococcus aureus (Staph. aureus). The primary clinical signs closely resembled a necrotizing fasciitis. Systemic and local application of vancomycin led to an improvement of the symptoms at secondary sites, but only the amputation of the primarily infected leg was lifesaving. CONCLUSION: In order to prevent such events, the authors recommend a number of additional presurgical measures. Firstly, a swab from the nose and throat should be taken prior to an elective surgery in patients with elevated risk of immunodeficiency, for example in patients with diabetes. The diabetes should be adequately treated before an elective operation is undertaken. Secondly, an early punction of the knee joint should be carried out if there is any doubt regarding inflammation. Isolated infectious agents should be grounds for early revision, which should always be completed with a rinsing procedure and with adequate antibiotics. Immunotherapy should be taken into consideration. Antiepidemic measures are recommended in cases with known Staph. aureus.

Aged↗

PubMed2003

Decrease of deleted in malignant brain tumour-1 (DMBT-1) expression is a crucial late event in intrahepatic cholangiocarcinoma.

AIMS: To investigate the participation of DMBT-1, a candidate tumour suppressor gene, in the development of intrahepatic cholangiocarcinoma via intraductal papillary neoplasm of the liver (IPN-L) arising in hepatolithiasis. DMBT-1 plays a role in mucosal immune defence. METHODS AND RESULTS: The expression of DMBT-1 was examined immunohistochemically in biliary epithelial cells in hepatolithiasis (n = 25), invasive and non-invasive cholangiocarcinoma associated with hepatolithiasis (n = 52), IPN-L with hepatolithiasis (n = 49), cholangiocarcinoma without hepatolithiasis (n = 32), and 10 normal control livers. DMBT-1 was expressed more frequently in the biliary epithelia of hepatolithiasis when compared with normal livers (P < 0.05). DMBT-1 expression was also frequent in IPN-L (57%) and non-invasive cholangiocarcinoma (79%). By contrast, DMBT-1 was decreased in invasive cholangiocarcinoma with and without hepatolithiasis (50% and 30%, respectively) (P < 0.05). The homozygous deletion of the DMBT-1 gene was recognized in four (20%) of 20 cholangiocarcinoma tissues and two (50%) of four cholangiocarcinoma cell lines, corresponding to the reduction of DMBT-1 expression. No deletion was detected in hepatolithiasis tissues. CONCLUSION: DMBT-1 expression is increased in IPN-L and non-invasive cholangiocarcinoma as well as in biliary epithelia in hepatolithiasis. Decreased expression of DMBT-1 and homozygous deletion of the DMBT-1 gene in invasive cholangiocarcinoma suggest that they occur in the late stage of cholangiocarcinogenesis.

Agglutinins↗

PubMed2002

Diffraction-enhanced X-ray imaging of articular cartilage.

OBJECTIVE: To introduce a novel X-ray technology, diffraction-enhanced X-ray imaging (DEI), in its early stages of development, for the imaging of articular cartilage. DESIGN: Disarticulated and/or intact human knee and talocrural joints displaying both undegenerated and degenerated articular cartilage were imaged with DEI. A series of three silicon crystals were used to produce a highly collimated monochromatic X-ray beam to achieve scatter-rejection at the microradian level. The third crystal (analyser) was set at different angles resulting in images displaying different characteristics. Once the diffraction enhanced (DE) images were obtained, they were compared to gross and histological examination. RESULTS: Articular cartilage in both disarticulated and intact joints could be visualized through DEI. For each specimen, DE images were reflective of their gross and histological appearance. For each different angle of the analyser crystal, there was a slight difference in appearance in the specimen image, with certain characteristics changing in their contrast intensity as the analyser angle changed. CONCLUSIONS: DEI is capable of imaging articular cartilage in disarticulated, as well as in intact joints. Gross cartilage defects, even at early stages of development, can be visualized due to a combination of high spatial resolution and detection of X-ray refraction, extinction and absorption patterns. Furthermore, DE images displaying contrast heterogeneities indicative of cartilage degeneration correspond to the degeneration detected by gross and histological examination.

Adult↗

PubMed2002

Analysis of losses of heterozygosity of the candidate tumour suppressor gene DMBT1 in melanoma resection specimens.

Deleted in malignant brain tumours 1 (DMBT1), a candidate tumour suppressor gene located on chromosome 10q25.3-q26.1, has recently been identified and found to be deleted in several different types of human tumours. In melanomas, the chromosomal region 10q22-qter is commonly affected by losses, hence we screened primary melanoma samples for losses of heterozygosity (LOH), and acquired melanocytic naevi and melanomas for transcription of DMBT1 and protein expression. Of 38 informative melanomas, 1 nodular melanoma and 2 subcutaneous metastases showed LOH of both microsatellites flanking the gene, suggesting loss of 1 DMBT1 allele. Three further melanomas showed LOH at 1 informative locus but were heterozygous for the second marker. Applying reverse-transcription polymerase chain reaction (RT-PCR), DMBT1 transcription was not found in melanomas. However, DMBT1 transcription was also absent from the majority of naevi from which melanomas frequently arise, making down-regulation of gene transcription during transformation from naevus to melanoma unlikely. Immunohistochemistry showed nerves, sweat glands and the stratum spinosum of the epidermis to be DMBT1 protein positive, whereas the naevi and melanoma cells themselves were negative. All considered, the candidate tumour suppressor gene DMBT1 does not appear to be a major inactivation target in the development of melanomas.

Agglutinins↗

PubMed2001

Deleted in Malignant Brain Tumors 1 is a versatile mucin-like molecule likely to play a differential role in digestive tract cancer.

Deleted in Malignant Brain Tumors 1 (DMBT1) has been proposed as a candidate tumor suppressor gene for brain, lung, and digestive tract cancer. In particular, alterations of the gene and/or a loss of expression have been observed in gastric, colorectal, and esophageal carcinomas. Initial evidence has accumulated that DMBT1 may represent a multifunctional protein. Because the consequences of a loss of DMBT1 function may be different depending on its original function in a particular tissue, we wondered if it is appropriate to assume a uniform role for DMBT1 in digestive tract carcinomas. We hypothesized that a systematic characterization of DMBT1 in the human alimentary tract would be useful to improve the understanding of this molecule and its role in digestive tract carcinomas. Our data indicate that the expression pattern and subcellular distribution of DMBT1 in the human alimentary tract is reminiscent of epithelial mucins. Bovine gallbladder mucin is identified as the DMBT1 homologue in cattle. An elaborate alternative splicing may generate a great variety of DMBT1 isoforms. Monolayered epithelia display transcripts of 6 kb and larger, and generally show a lumenal secretion of DMBT1 indicating a role in mucosal protection. The esophagus is the only tissue displaying an additional smaller transcript of approximately 5 kb. The stratified squamous epithelium of the esophagus is the only epithelium showing a constitutive targeting of DMBT1 to the extracellular matrix (ECM) suggestive of a role in epithelial differentiation. Squamous cell carcinomas of the esophagus show an early loss of DMBT1 expression. In contrast, adenocarcinomas of the esophagus commonly maintain higher DMBT1 expression levels. However, presumably subsequent to a transition from the lumenal secretion to a targeting to the ECM, a loss of DMBT1 expression also takes place in adenocarcinomas. Regarding DMBT1 as a mucin-like molecule is a new perspective that is instructive for its functions and its role in cancer. We conclude that DMBT1 is likely to play a differential role in the genesis of digestive tract carcinomas. However, although DMBT1 originally has divergent functions in monolayered and multilayered epithelia, carcinogenesis possibly converges in a common pathway that requires an inactivation of its functions in the ECM.

Adenocarcinoma↗

PubMed2001

Myeloid-related protein (MRP)-8 from cervico-vaginal secretions activates HIV replication.

OBJECTIVES: To identify a substance found in female genital tract secretions that enhances HIV expression in infected cells. DESIGN: Cervico-vaginal lavages (CVL), collected in sterile normal saline, were fractionated and tested for HIV-inducing activity using HIV-infected monocytes. METHODS: To purify the component(s) of CVL that enhance HIV production, Mono-Q ion exchange chromatography followed by Superose-12 molecular sieve analysis, and SDS--PAGE were performed. The purified protein was identified by amino acid sequence analysis. RESULTS: SDS--PAGE of bioactive fractions showed a 14 kDa polypeptide band. Amino acid sequence analysis of selected peptides from the 14 kDa band showed 100% homology with the myeloid-related protein (MRP)-8, an inflammatory protein found in mucosal secretions. Western blot analysis revealed that bioactive CVL contained more immunoreactive MRP-8 than samples without bioactivity. The HIV-inducing activity of MRP-8 was further confirmed by showing that human recombinant MRP-8 increased HIV expression by up to 40-fold. CONCLUSIONS: MRP-8 in cervico-vaginal secretions stimulates HIV production. Strategies aimed at blocking MRP-8 activity in the genital tract could reduce risk of sexual as well as maternal--infant transmission of HIV.

Amino Acid Sequence↗

PubMed2001

Allelic imbalance and fine mapping of the 17p13.3 subregion in sporadic breast carcinomas.

Chromosome arm 17p is frequently altered in a variety of human cancers, especially in breast cancer, and allelic imbalances (AIs) in the region 17p13.1 do not always coincide with mutations in the TP53 gene. A second interval that frequently shows AIs at 17p is the chromosomal band 17p13.3. This region is suspected to harbor another tumor suppressor gene. In order to get more information concerning the pattern of AIs in 17p13.3, we performed analysis of AI of 49 breast carcinomas at 6 polymorphic loci in 17p13.3. Eighty-six percent of the tumors present AI at least at one marker in 17p13.3. Among all loci tested, the highest percentage of Al was observed at loci D17S5 (77%) and D17S1528 (72%). According to these results, a minimal region of deletion could be determined between the markers D17S28 and D17S5. Fine mapping of this region revealed that the size of the deleted region is about 100-150 kb. Furthermore, a subset of the patients shows two other areas with AI close to the markers D17S1574/D17S513 and D17S849, respectively.

Allelic Imbalance↗

PubMed2001

Mechanisms of chondrocyte adhesion to cartilage: role of beta1-integrins, CD44, and annexin V.

The initial adhesion of transplanted chondrocytes to surrounding host cartilage may be important in the repair of articular defects. Adhesion may position cells to secrete molecules that fill the defect and integrate repair tissue with host tissue. While chondrocytes are known to become increasingly adherent to cartilage with time, the molecular basis for this is unknown. The objective of this study was to investigate the role of beta1-integrin, CD44, and annexin V receptors in chondrocyte adhesion to cartilage. Chondrocytes were cultured in high density monolayer, released with trypsin, and allowed to recover in suspension for 2 h at 37 degrees C. Under these conditions, flow cytometry analysis showed that chondrocytes expressed beta1-integrins, CD44, and annexin V. In a rapid screening assay to assess chondrocyte adhesion to cartilage, cell detachment decreased from 79% at 10 min following transplantation to 10% at 320 min. Treatment of cells with a monoclonal antibody to block beta1-integrins significantly increased chondrocyte detachment from cartilage compared to untreated controls. Similarly, results from a parallel-plate shear flow adhesion assay showed that blocking beta1-integrins significantly increased chondrocyte detachment from cartilage compared to untreated controls at each level of applied shear (0-70 Pa). In both assays, treatment of cells with reagents that block CD44 (hyaluronan oligosaccharides or monoclonal Ab IM7) or annexin V (polyclonal Ab #8958) had no detectable effect on adhesion. With cartilage treated with chondroitinase ABC, blocking beta1-integrins also increased chondrocyte detachment, while blocking CD44 and annexin V also had no detectable effect. Under the conditions studied here, beta1-integrins appear to mediate chondrocyte adhesion to a cut cartilage surface. Delineation of the mechanisms of adhesion may have clinical implications by allowing cell manipulations or matrix treatments to enhance chondrocyte adhesion and retention at a defect site.

Animals↗

PubMed2001

Donor fascia in urogynaecological procedures: a canine model.

OBJECTIVE: To explore the in vivo characteristics of donor fascia used in urogynaecological procedures, in a canine model. MATERIALS AND METHODS: Two experiments were conducted. In the first, donor fascia grafts were obtained from 12 dogs, the grafts freeze-dried and half were irradiated. The grafts were used for sacrocolpopexy and suburethral slings in each of five dogs. The dogs were killed at 2, 6 and 12 weeks after graft implantation, the grafts retrieved and assessed using tensilometry. In the second experiment, unirradiated sacrocolpopexy grafts were implanted in eight dogs; four grafts were placed under no tension and four under moderate tension. At 8 weeks, the grafts were retrieved and assessed by tensilometry. Measures of strength in both experiments included the ultimate tensile strength, ultimate strain and stiffness. All measures were compared using Kruskal-Wallis nonparametric tests in both studies. RESULTS: In the first experiment, a significant minority (23%) of grafts had complete loss of strength. Measures of graft strength did not vary when analysed according to donor dog, host dog, history of graft irradiation, duration of implantation or location of graft. In the second experiment, grafts placed under no tension tended to have lower tensile strength (chi2(1) = 3.125, P = 0.077), lower stiffness (chi2(1) = 3.125, P = 0.077) and lower ultimate strain (chi 2(1) = 3.182, P = 0.074). CONCLUSION: Graft irradiation as an isolated variable did not predispose grafts to failure in vivo. Biomechanical factors at the implantation site are likely to play a critical role in determining ultimate graft strength.

Animals↗

PubMed2001

The effects of collagen fragments on the extracellular matrix metabolism of bovine and human chondrocytes.

Cartilage matrix degradation generates collagen type II fragments. The objective of this study is to explore the possibility that these collagen fragments may be part of an endogenous metabolic feedback. Initially, collagen fragments were extracted from normal or osteoarthritic cartilage, as part of a matrix fragment preparation. Later, collagen fragments were generated by digestion of bovine collagen type II with bacterial collagenase (col2f). These fragments were added to cultures of isolated chondrocytes (bovine and human) and cartilage explants (human). In a dose-dependent manner, col2f caused inhibition of cell attachment to collagen, inhibition of collagen synthesis, and induction of matrix degradation. In addition, when col2f were added to human cartilage explants, an induction of gelatinase activity was detected in the media. These data sets present first evidence that degradation products of collagen may be directly involved in the regulation of cartilage homeostasis.

Adult↗

PubMed2000

Physical and transcriptional mapping of the 17p13.3 region that is frequently deleted in human cancer.

Studies of chromosomal losses at 17p13 have suggested the presence of at least two distinct regions for tumor suppressor genes, the TP53 region at 17p13.1 and a more distal region at 17p13.3. Within the latter region, Hypermethylated in Cancer 1 (HIC1) is located, a likely candidate for a tumor suppressor gene that has also been suggested to play a role in the pathogenesis of Miller-Diecker syndrome (MDS). However, single-gene isolation efforts have retrieved additional genes from 17p13.3 that could play a role in tumorigenesis. This indicates that the full potential of this chromosomal region with respect to disease-related genes has not yet been exhausted and that there may exist still unknown genes that contribute to tumorigenesis or to the complex MDS phenotype. To provide a basis for the systematic isolation and evaluation of such genes, we established a physical map over 1.5 Mb of 17p13.3 and assigned 29 transcriptional units within this region.

Abnormalities, Multiple↗