Visualization of bone formation in sheep's middle ear by using fluorochrome sequential labelling (FSL).

One factor for the lacking integration of the middle ear stapes footplate prosthesis or the missing healing of stapes footplate fractures could be the known osteogenic inactivity. In contrast, it was recently demonstrated that titanium prostheses with an applied collagen matrix and immobilised growth factors stimulate osteoblastic activation and differentiation on the stapes footplate. Regarding those findings, the aim of this study was to evaluate the potential of bone regeneration including bone remodeling in the middle ear. Ten one-year-old female merino sheep underwent a middle ear surgery without implantation of middle ear prostheses or any other component for activating bone formation. Post-operatively, four fluorochromes (tetracycline, alizarin complexion, calcein green and xylenol orange) were administered by subcutaneous injection at different time points after surgery (1 day: tetracycline, 7 days: alizarin, 14 days: calcein, 28 days: xylenol). After 12 weeks, the temporal bones including the lateral skull base were extracted and histologically analyzed. Fluorescence microscopy analysis of the entire stapes with the oval niche, but in particular stapes footplate and the Crura stapedis revealed evidence of new bone formation. Calcein was detected in all and xylenol in 60% of the animals. In contrast, tetracycline and alizarin could only be verified in two animals. The authors were able to demonstrate the osseoregenerative potential of the middle ear, in particular of the stapes footplate, using fluorescence sequence labelling.

Satellite cell capacity for functional adaptation of masseter muscle in Class II and Class III patients after orthognathic surgery-a pilot study.

AIM: The aim of the prospective pilot study was to analyze the biomarkers CD34, Pax7, Myf5, and MyoD for stimulation of satellite cells (SCs), which are responsible for functional adaptation. SUBJECTS AND METHODS: Forty-five Caucasian patients were consecutively recruited from the Maxillo-Facial-Surgery at TU Dresden. Eleven orthognathic Class III patients, 24 Class II patients, and 10 controls with Class I were involved in the study. Tissue samples from masseter muscle were taken from the patients pre-surgically (T1) and 7 months later (T2). Samples from controls were taken during the extraction of third molars in the mandible. Polymerase chain reaction (PCR) for relative quantification of gene expression was calculated with the delta delta cycle threshold (ΔΔCT) method. RESULTS: The results show significant differences for the marker of SC stimulation between the controls, the patient groups, males, and females. The gene expression of CD34 was post-surgically upregulated for Class III (0.35-0.77, standard deviation [SD] = 0.39, P < 0.05) in comparison with controls. For Pax7, there was a significant difference shown between the retrognathic and the prognathic group because of downregulation in Class II patients (1.64-0.76, SD = 0.55, P < 0.05). In Class III patients, there was a significant upregulation for Myf5 (0.56-1.05, SD = 0.52, P < 0.05) after surgery too. CONCLUSIONS: The significant decline of Pax7 in Class II patients indicates a deficiency of stimulated SC post-surgically. The expression of CD34 and Myf5 in Class II stayed unchanged. In contrast, there was an upregulation for all Class III patients, mainly in females, shown post-surgically. This may be one reason for weak functional adaptation and relapse in Class II patients.

Notch expression profile and satellite cell stimulation in masseter muscle before and after orthognathic surgery.

The aim of this prospective study was to compare the expression of the Notch receptor family with the biomarker for stimulation of satellite cells (SC), which are responsible for functional adaptation. Tissue samples from the masseter muscle were taken presurgically and 7 months later. Samples from controls came from the extraction of third molars. The expression of Notch 1 to 4 and the satellite cell markers CD34, Pax7, and MyoD1 were investigated. PCR was used for relative quantification of gene expression, which was calculated with the ΔΔCT method. The study involved 38 white patients - 10 prognathic, 18 retrognathic, and 10 orthognathic controls. The median value for Notch 1 was significantly reduced presurgically for prognathic (0.46, SD 0.45) and retrognathic (0.57, SD 0.35) patients compared with the controls. Postsurgically, Notch 2 was significantly upregulated in the prognathic group (0.55, SD 0.28/1.37, SD 0.85). Similarly, there was upregulation of Notch 3 in the prognathic group (0.33, SD 0.42/0.59, SD 1.37) and downregulation in retrognathic patients (0.59, SD 0.79/0.52, SD 0.97). Upregulations for the satellite cell markers CD34 and Pax7 were also found in prognathic patients. The significant upregulation of Notch 1-3 and CD34 in prognathics, but unchanged MyoD expression, signals high stimulation for SC and maintenance of the regeneration cell pool. A lower expression of Notch and SC in retrognathic patients could be responsible for weak functional adaptation.

The Obliteration of Noncritical Size Bone Defects With Bone Dust or Bone Replacement Material (Bioactive Glass S53P4).

HYPOTHESIS: Bone dust (BD) harvested during operation may be suitable as an autologous obliteration material for noncritical size defects. Bioactive glass (BA) can be an alternative. BACKGROUND: To treat noncritical size defects, BD and BA are commonly used for obliteration techniques. However, the optimal harvesting method and parameters for BD have not been examined. In this study, we analyzed the osseoregenerative potential of both materials. METHODS: Thirteen female merino sheep (7-yr old) underwent surgery on the frontal calvaria. Three defects were inserted. The first defect was considered a reference and remained unfilled, the second defect was filled with BD from the calvaria bone, and the third defect was filled with BA S53P4. The animals were sacrificed after 3 weeks. To evaluate bone regeneration, we used digital volume tomography, bone density measurement, fluorochrome sequence labeling, and histological analysis. RESULTS: All analyses showed quantitative and qualitative bone regeneration 3 weeks after operation. The control blank defect showed significantly less new bone growth than the BD-filled defect. Moreover, bone regeneration occurred from the surrounding bone and showed only a defect bridge in the BD-filled defect. The BA completely filled the defect and had the highest density although the same amount of new mineralized bone generated as in the reference. CONCLUSION: BD and BA seemed to be suitable bone replacement materials for obliteration techniques because they completely filled the defects. Thus, BD harvested under standardized conditions provided a higher level of osteoreparation potential for the generation of woven bone and establishment of defect bridges.

Histological examinations of the in vivo biocompatibility of oxycellulose implanted into rat skeletal muscle.

BACKGROUND: Recently it was shown that oxycellulose suppressed bone regeneration led to an accumulation of connective tissue as well as foam cells in bone defects. OBJECTIVES: Since oxycellulose can be used as a hemostatic agent in general and dental surgery, the aim of the study was to examine muscle tissue response to implanted oxidized cellulose. MATERIAL AND METHODS: RESO-Cell® (Resorba Wundversorgung GmbH, Nuremberg, Germany) standard was implanted in the latissimus dorsi of 20 rats; subsequently, 12 samples were processed for histological evaluation after 4 and 8 weeks. The remaining 8 samples were processed for mRNA expression analyses of gene-encoding growth factors and collagens using quantitative reverse transcription polymerase chain reaction (RT-qPCR). RESULTS: Muscle tissue exposed to oxycellulose showed elevated mRNA levels of COL1A1 compared to untreated muscle tissue. The histological analysis revealed that no undegraded oxycellulose was detectable after as little as 4 weeks. Furthermore, a strong accumulation of CD68-positive foam cells was found in the treated area. CONCLUSIONS: In conclusion, the study has shown that oxidized cellulose can cause an inflammatory response after this material is implanted in skeletal muscle. Therefore, it is not recommended to leave this material in the body over a long period. However, it could be used as auxiliary material in the treatment of periodontal defects.

Interrelationship between bone substitution materials and skeletal muscle tissue.

Bone density and quantity are primary conditions for the insertion and stability of dental implants. In cases of a lack of adequate maxillary or mandibulary bone, bone augmentation will be necessary. The use of synthetic bioactive bone substitution materials is of increasing importance as alternatives to autogenously bone grafts. It is well known that bone can influence muscle function and muscle function can influence bone structures. Muscles have a considerable potential of adaptation and muscle tissue surrounding an inserted implant or bone surrogate can integrate changes in mechanical load of the muscle and hereupon induce signaling cascades with protein synthesis and arrangement of the cytoskeleton. The Musculus latissimus dorsi is very often used for the analyses of the in vivo biocompatibility of newly designed biomaterials. Beside macroscopically and histologically examination, biocompatibility can be assessed by analyses of the biomaterial influence of gene expression. This review discusses changes in the fiber type distribution, myosin heavy chain isoform composition, histological appearance and vascularization of the skeletal muscle after implantation of bone substitution materials. Especially, the effects of bone surrogates should be described at the molecular-biological and cellular level.

Histological and molecular-biological analyses of poly(3-hydroxybutyrate) (PHB) patches for enhancement of bone regeneration.

Tissue engineered cell-seeded constructs with poly(3)hydroxybutyrate (PHB) induced ectopic bone formation after implantation into the back muscle of rats. The objective of our in vivo study was to evaluate the osteogenic potential of pure PHB patches in surgically created cranial defects. For this, PHB patches were analyzed after implantation in surgically created defects on the cranium of adult male rats. After healing periods of 4, 8 and 12 weeks, the bone tissue specimens containing PHB patches were processed and analyzed histologically as well as molecular-biologically. After 4 weeks, the PHB patches were completely embedded in connective tissue. Eight weeks after PHB insertion, bone regeneration proceeding from bearing bone was found in 50% of all treated animals, whereas all PHB treated cavities showed both bone formation and embedding of the patches in bone 12 weeks after surgery. Furthermore, all slices showed pronounced development of blood vessels. Histomorphometric analysis presented a regenerated bone mean value between 46.4 ± 16.1% and 54.2 ± 19.3% after 4-12 weeks of healing. Caveolin-1 staining in capillary-like structures showed a 1.16-1.38 fold increased expression in PHB treated defects compared to controls. Real-time RT-PCR analyses showed significantly lower expressions of Alpl, Col1a1 and VEGFA in cranium defects after treatment with PHB patches compared to untreated bony defects of the same cranium. Within the limits of the presented animal investigation, it could conclude that the tested PHB patches featured a good biocompatibility and an osteoconductive character.

Immunosuppressive Glycodelin A is an independent marker for poor prognosis in endometrial cancer.

BACKGROUND: Knowledge on immunosuppressive factors in the pathogenesis of endometrial cancer is scarce. The aim of this study was to assess Glycodelin (Gd) and its immunosuppressive isoform Glycodelin A (GdA) in endometrial cancer tissue and to analyze its impact on clinical and pathological features and patient outcome. METHODS: 292 patients diagnosed and treated for endometrial cancer were included. Patient characteristics, histology and follow-up data were available. Gd and GdA was determined by immunohistochemistry and in situ hybridization was performed for Gd mRNA. RESULTS: Endometrial cancer shows intermediate (52.2%) or high (20.6%) expression for Gd in 72.8%, and GdA in 71.6% (intermediate 62.6%, high 9.0%) of all cases. The glycosylation dependent staining of GdA is tumour specific and correlates with the peptide-specific Gd staining though neither of the two is associated with estrogen receptor, progesterone receptor or clinic-pathological features. Also Gd protein positively correlates with Gd mRNA as quantified by in situ hybridization. Gd positive cases have a favourable prognosis (p = 0.039), while GdA positive patients have a poor outcome (p = 0.003). Cox-regression analysis proofed GdA to be an independent prognostic marker for patient survival (p = 0.002), besides tumour stage, grade and the concomitant diagnosis of hypertension. CONCLUSION: Gd and GdA are commonly expressed in endometrial cancer tissue and seem to be of relevance in tumourigenesis. They differ not only in glycosylation but also in their biological activity, since only GdA holds prognostic significance for a poor overall survival in endometrial cancer patients. This finding might be explained by GdAs immunosuppressive capacity.

Histological evaluation at different times after augmentation of extraction sites grafted with a magnesium-enriched hydroxyapatite: double-blinded randomized controlled trial.

OBJECTIVES: To histologically analyze the early angiogenesis-osteogenesis interplay in post-extraction sockets augmented with magnesium-enriched hydroxyapatite (Mg-enriched HA). MATERIAL AND METHODS: Ten post-extraction sites underwent post-extraction ridge preservation procedure. According to randomization, sites were divided into two balanced groups and bone specimens were collected 2 or 4 months after surgery. Sections were stained with hematoxylin/eosin, Masson-Goldner trichrome, and tartrate-resistant acid phosphatase (TRAP), respectively. Furthermore, indirect immunohistochemistry was performed using alkaline phosphatase, CD34 and caveolin-1 antibodies. Mean values and standard deviations were calculated for each outcome variable. Data were then compared using one-way ANOVA test. P < 0.05 was considered statistically significant. RESULTS: Histomorphometric analysis presented 15.0% (±3.5) regenerated bone after 2 months of healing. After 4 months, regenerated bone increased 5.1-fold up to 77.4% (± 8.6) (P < 0.001). On the contrary, vessels and capillary reduced from 645 (±33) to 255 (± 94) (caveolin-1 expression, P = 0.008). These findings were confirmed by CD34 expression (301 ± 95 and 88 (±24), respectively, at 2 and 4 months (P = 0.046). CONCLUSIONS: Within the limits of the present randomized controlled study, it can be concluded that Mg-enriched HA is a suitable material for socket preservation and ensures early angiogenesis and early osteogenesis.

Bone substitution materials on the basis of BONITmatrix® up-regulate mRNA expression of IGF1 and Col1a1.

The aim of this study was to investigate the effects of BONITmatrix(®) and OSSA NOVA on the expression of growth factors and osteogenic differentiation. For this purpose, the mRNA expression of VEGF, IGF1, IGF2, collagen-1, collagen-2 and MMP8 was analysed in surgically created defects on the crania of adult male rats. Cranial samples were collected after implantation of BONITmatrix(®) or OSSA NOVA scaffolds for 4 weeks and determinations of gene expression were performed by quantitative RT-PCR. Real-time RT-PCR analyses showed a significantly higher expression of IGF1 in both groups treated with BONITmatrix(®) and OSSA NOVA compared to untreated controls, whereas type I collagen mRNA expression only increased in BONITmatrix(®) treated rats compared to controls. No changes in transcript expression of IGF2, VEGF, collagen-2 and MMP8 were detectable between the analysed groups. In conclusion, BONITmatrix(®) and OSSA NOVA stimulate the expression of growth factor IGF1, but only the granular dosage form is able to stimulate osteoblast differentiation.

Experimental and histological investigations of the bone using two different oscillating osteotomy techniques compared with conventional rotary osteotomy.

Over the past decade, coinciding with the appearance of a number of new ultrasonic surgical devices, there has been a marked increase in interest in the use of ultrasound in oral surgery and implantology as alternative osteotomy method. The aim of this study was the comparison of the effect of osteotomies performed using ultrasonic surgery (Piezosurgery(®)), sonic surgery SONICflex(®) and the conventional bur method on the heat generation within the bone underneath the osteotomy and light-microscopy observations of the bone at different cutting positions in porcine mandibular segments. It was found that the average heat generated by SONICflex(®) sonic device was close to that by conventional rotary bur (1.54-2.29°C), whereas Piezosurgery(®) showed a high generated heat up to 18.17°C. Histological investigations of the bone matrix adjacent to the defect radius showed intact osteocytes with all three instruments and similar wide damage diameter at the bottom region. SONICflex(®) showed smooth cutting surfaces with minimal damage in the upper defect zone. Finally, presented results showed that sonic surgery performed with SONICflex(®) is an alternative osteotomy method and can be used as an alternative to the conventional bur method.

Changes in condylar cartilage after anterior mandibular displacement in juvenile pigs.

UNLABELLED: Adaptive remodelling of the mandibular condyle in response to mandibular advancement is the mechanism exploited by orthodontic forward displacement devices. OBJECTIVE: This work investigated the expression of collagens, matrix metalloproteinases and vascular endothelial growth factor during this process. DESIGN: Twenty juvenile pigs were randomly divided into two experimental groups, where the treatment group was fitted with mandibular advancement splints, and the control group was not. Changes in the mRNA content of condylar cartilage tissue was then were measured by real-time PCR using specific primers after 4weeks of treatment. RESULTS: The temporal pattern of the expression of Col1 and MMP13 during condylar adaptation coincided with that during natural condylar growth. The amount of the expression of Col10 during condylar adaptation was significantly lower (p<0.05), whereas the expression of Col2, MMP8 and VEGF was significantly higher compared to natural growth (p<0.05). CONCLUSIONS: It is suggested that condylar adaptation in growing pigs triggered by mandibular forward positioning results not only from passive adaptation of cartilage, but also involves growth affected processes. Our results showed that mechanical strain produced by mandibular advancement induced remodelling and revascularization in the posteriocranial mandibular condyle. These results are mostly consistent with former published histological and histomorphometrical analyses.

Caveolin-1, caveolin-3 and VEGF expression in the masticatory muscles of mdx mice.

Duchenne muscular dystrophy (DMD) and murine X-linked muscular dystrophy (mdx), its murine model, are characterized by muscle damage and muscle weakness associated with inflammation and new vessel formation. Caveolins, dystrophin-associated proteins, are involved in the pathogenesis of DMD, because increased numbers of caveolae are found in DMD and mdx hindlimb muscles. Caveolae influence angiogenesis due to their content of vascular endothelial growth factor (VEGF) receptors. Orofacial muscles in mdx mice undergo muscle necrosis followed by muscle regeneration. To ascertain the role of caveolins and VEGF in the pathogenesis of dystrophic masticatory muscles, we examined the expression of caveolin-1 (cav-1), caveolin-3 (cav-3) and VEGF in control and mdx mice. In mdx masticatory muscles, no changes in transcript and protein levels of VEGF were found, whereas cav-1 and cav-3 expression was increased. Using immunohistochemistry, a strong sarcolemmal staining of caveolin-3 in regenerated muscle fibers was found. Furthermore, immunohistochemistry with the caveolin-1 antibody showed an increase in the amount of blood vessels in areas with regenerating muscle fibers. Dystrophic masticatory muscles showed changes comparable to those of hindlimb muscles in the expression of cav-1 and cav-3. The angiogenesis seems to be unaffected in the jaw muscles of mdx mice. We speculate that the increased caveolin expression could cause extensive and efficient muscle regeneration.

Immunolocalization of glycodelin in human adenocarcinoma of the lung, squamous cell carcinoma of the lung and lung metastases of colonic adenocarcinoma.

Glycodelin (Gd), which is localized in cells of bronchial epithelium, type II pneumocytes and alveolar macrophages in rats and humans, plays an important role in the pulmonary immune response in asthmatic inflammation. In this study, sections of paraffin-embedded tumor adjacent lung tissue and sections of adenocarcinoma of the lung, squamous cell carcinoma of the lung and metastases of colonic adenocarcinoma were investigated for the distribution and expression of Gd using a polyclonal anti-Gd antibody. Glycodelin protein is located in the cytoplasm of bronchial epithelial cells, pneumocytes and alveolar macrophages. Furthermore, Gd is expressed in adenocarcinoma and squamous cell carcinoma of the lung as well as in lung metastases of colonic adenocarcinoma. Densitometric analyses showed a significantly increased expression of glycodelin protein in cancer tissue compared to tumor adjacent lung tissue. The Gd protein level was 1.7-2.6-fold increased in lung carcinoma compared to tumor adjacent lung tissue. The Gd protein level did not differ from each other between the investigated types of cancer tissue. Because these data validate the recent findings of Gd mRNA expression, it may be concluded that glycodelin plays an important role in the pathogenesis of lung cancer and lung metastases.

Immunohistochemistry, glycosylation and immunosuppression of glycodelin in human ovarian cancer.

Glycodelins (Gds) are glycoproteins with a gender specific glycosylation. Glycodelin A (GdA) is primarily produced in endometrial and decidual tissue and secreted to amniotic fluid. Glycodelins were also identified in several cancer types, including serous ovarian cancer. Gds act as a T-cell inhibitor and are involved in inactivation of human monocytes. With a Gd peptide antibody, derived from a 15 amino acid sequence of human Gd and in situ hybridization experiments, the expression of Gd in serous, mucinous, endometrioid and clear cell ovarian tumors was identified. In contrast to former investigations with antibodies against GdA, a positive immunohistochemical reaction for Gd was observed in all forms of epithelium ovarian cancer. These results were confirmed with in situ hybridization. In addition, Gd is expressed in granulose cell tumors, a non-epithelial form of ovarian cancer. Furthermore, Gd was purified from ascites fluid of ovarian cancer patients. Ascites Gd showed significant differences in its structure of sialyl Lewis-type oligosaccharides compared to GdA. Additionally, ascites Gd inhibits IL-2 stimulated proliferation of peripheral blood leucocytes and inhibits adhesion of SLe(X)-positive cells to E-selectin. Therefore, Gd could act as an inhibitor of lymphocyte activation and/or adhesion in ovarian cancer.

Glycodelin protein and mRNA is downregulated in human first trimester abortion and partially upregulated in mole pregnancy.

Glycodelin (Gd) is a major reproductive glycoprotein and a mediator for immunomodulatory effects directed to cellular, humoral, and innate immunity. Human pregnancy depends on a diversity of physiological processes including modulation of the maternal immunosystem. We evaluated the expression of Gd protein and mRNA in first trimester decidual tissue of normal pregnancies and spontaneous abortion and hydatidiform moles. Furthermore, in vitro experiments on endometrial cancer cells to analyze the effect of human chorionic gonadotropin (hCG) on Gd regulation were performed. In decidual tissue of abortion patients, Gd expression was significantly decreased compared with normal gestation, which was confirmed by in situ hybridization. In mole pregnancy, an upregulation of Gd in the first 8 weeks of pregnancy was present. Gd is a main product of decidual tissue in the first trimester of human pregnancy. Reduced Gd expression in abortive pregnancy could lead to an increased activation of the maternal immunosystem, thus causing rejection of the developing fetus. Moreover, Gd expression in endometrial cancer cells in vitro could be stimulated by addition of hCG. Therefore, we speculate that hCG could be one of the factors regulating Gd expression because hCG is downregulated in women with abortion and upregulated in mole pregnancy. In addition, we found a positive feedback loop in Gd and hCG expression in human pregnancy.

Expression of E-cadherin in human ductal breast cancer carcinoma in situ, invasive carcinomas, their lymph node metastases, their distant metastases, carcinomas with recurrence and in recurrence.

BACKGROUND: Breast cancer cells can invade and generate metastasis via either lymphatic or blood vessels. E-cadherin mediates tumor cell-cell adhesion. Partial or complete loss of E-cadherin expression correlates with poor prognosis in breast cancer patients. In this study, the expression of E-cadherin was examined in mammary ductal carcinoma in situ, invasive breast carcinomas without metastasis, invasive carcinomas with their lymph node and distant metastases and invasive carcinomas with local recurrence in breast cancer tissue. MATERIALS AND METHODS: Paraffin-embedded slides of carcinoma in situ (8 DCIS), invasive carcinomas without lymph node metastases (9 invasive ductal carcinomas), invasive carcinomas (7 invasive ductal carcinomas) with corresponding lymph node metastases, invasive carcinomas (8 invasive ductal carcinomas) with corresponding recurrence and invasive carcinomas (5 invasive ductal carcinomas) with corresponding distant metastases were investigated for E-cadherin expression. Tissue slides were incubated with monoclonal antibodies against E-cadherin and stained with the ABC-elite kit. Staining intensities were analyzed by using a semi-quantitative score. RESULTS: A strong expression of E-cadherin in carcinoma in situ was demonstrated. Expression of E-cadherin was moderate in invasive carcinomas without metastases. However, very weak expression of E-cadherin in primary carcinoma with lymph node metastases was detected. E-cadherin expression was elevated in lymph node metastases compared to the primary tumor. CONCLUSION: Analysis of a tumor antigen involved in adhesion of breast cancer cells showed that there are significant differences of expression of E-cadherin between primary breast cancer cells and their metastases. Evaluation of this marker involved in cell adhesion could be a useful method for evaluating the metastatic risk in breast cancer patients.

Glycodelin A is expressed differentially in normal human endometrial tissue throughout the menstrual cycle as assessed by immunohistochemistry and in situ hybridization.

OBJECTIVE: To [1] evaluate glycodelin A immunolabeling in normal endometrium with specific monoclonal (mAb) and polyclonal peptide (pAb) antibodies, [2] to assess glycodelin messenger RNA (mRNA) by in situ hybridization, and [3] to conduct deglycosylation experiments to evaluate the recognized epitope of the mAb vs. pAb. DESIGN: Retrospective immunohistochemical analysis. SETTING: University institute and hospital in Germany. PATIENT(S): Normal human endometrial tissue from the proliferative (PP), early secretory, and late secretory phases were obtained from patients undergoing surgery for benign diseases. INTERVENTION(S): Generation of a pAb in rabbit, immunohistochemistry, and in situ hybridization. MAIN OUTCOME MEASURE(S): Semiquantitative and computerized analysis. RESULT(S): A statistically significant increase of the glycodelin A immunolabeling in the late secretory phase compared with PP was demonstrated by using the mAb. Polyclonal-peptide antibody immunolabeling also showed a rise between the PP and late secretory phases, but without statistical significance. In situ hybridization demonstrated a statistically significantly higher mRNA content during the early secretory phase compared with during PP. CONCLUSION(S): Glycodelin was demonstrated in normal endometrium at the protein and mRNA levels. The mAb may be more useful in assessing glycodelin expression in endometrium, because it probably can bind to glycodelin A-unique glycan structures, in contrast to the peptide pAb. This is of major interest because it may reveal possible structural and functional relationships in different parts of this molecule and elucidate possible functions of this glycoprotein in human endometrial tissue.

MDR1 gene polymorphisms and risk of gingival hyperplasia induced by calcium antagonists.

BACKGROUND: Gingival overgrowth is a common side effect of calcium antagonists. Although the pathogenesis is unknown, several lines of evidence point to a modulation of inflammatory processes. Because the calcium antagonists, albeit to a variable degree, act as inhibitors of P-glycoprotein (P-gp), the gene product of multidrug resistance 1 (MDR1), and inflammation may modify P-gp expression, we analyzed the MDR1 polymorphisms as risk factors for gingival overgrowth induced by calcium antagonists. METHODS: Clinical, laboratory, and anamnestic data including periodontal parameters and use of calcium antagonists were assessed in a cross-sectional epidemiologic investigation (N = 1484). MDR1 polymorphisms in exon 21 G2677T/A and exon 26 C3435T were determined. P-gp expression was detected in gingival tissues. In a matched-pair analysis, 93 subjects using calcium antagonists and 186 not using them were compared. RESULTS: P-gp is expressed in the endothelial layers of blood vessels obtained from healthy or inflamed gingiva. Subjects treated with calcium antagonists had significantly deeper gingival pockets than their drug-free counterparts (P <.0001). This drug-related side effect was associated with the MDR1 2677G/G or G/TA genotype (P <.001) but not with the variant genotype T/TA. This drug effect was proved by multiple regression analysis with adjustment for the risk factors of periodontitis (age, sex, smoking, and education) (P <.0001) and was associated with elevated C-reactive protein levels. The association of probing depth with the MDR1 polymorphism was confirmed in the matched-pair analysis (P <.0001). CONCLUSION: Treatment with calcium antagonists leads to gingival hyperplasia, which is associated with the MDR1 G2677T/A polymorphism. The MDR1 genotype may modify the inflammatory response to the drugs.

Expression of sialyl lewis X, sialyl Lewis A, E-cadherin and cathepsin-D in human breast cancer: immunohistochemical analysis in mammary carcinoma in situ, invasive carcinomas and their lymph node metastasis.

OBJECTIVE: Breast cancer cells can invade and generate metastasis via either lymphatic or blood vessels. Sialyl Lewis X (SLeX) and Sialyl Lewis a (SLea) are carbohydrate molecules that mediate the adhesion between tumor cells and the endothelium. These antigens are not expressed on normal breast tissue. Overexpression of SLeX and SLea is combined with poor prognosis and malignant relapse. E-cadherin mediates tumor cell-tumor cell adhesion. Partial or complete loss of E-cadherin expression has also been found to correlate with poor prognosis in breast cancer patients. Another factor involved in metastasis is Cathepsin-D, a lysosomalprotease. Cathepsin-D plays a role in cell proliferation and inhibition of tumor cell adhesion. In this study, we analysed the combined expression of SLeX, SLea, Cathepsin-D and E-cadherin in breast carcinoma in situ, invasive carcinomas without metastasis and invasive carcinomas with lymph node metastasis. MATERIALS AND METHODS: Slides of paraffin-embedded tissue of carcinoma in situ (8 DCIS, 2 CLIS), invasive carcinomas without metastasis (9 ductal, 1 lobular) and carcinomas (7 ductal, 2 lobular, 1 mucinous) with their lymph node metastasis (10 cases) were used. Breast cancer tissue was fixed and incubated with monoclonal antibodies against SLex (IgM) and SLea (IgM), Cathepsin-D (IgG) and E-cadherin (IgG). Staining reaction was performed with the ABC reagent. The intensity of immunohistochemical reaction on digital images of the slides was analyzed using a computer-aided detection system. RESULTS: We found a weak expression of both Sialyl Lewis antigens, a strong expression of E-cadherin and a moderate expression of Cathepsin-D in carcinoma in situ. The expression of both Sialyl Lewis antigens, E-cadherin and Cathepsin-D was moderate in invasive carcinomas without metastases. However, a strong expression of both Sialyl Lewis antigens and a very weak expression of E-cadherin was detected in primary carcinoma with lymph node metastases. The expression of Cathepsin-D was moderate in this tissue. E-cadherin expression was elevated whereas SLeX and Cathepsin-D expression was reduced in lymph node metastases compared to the primary tumor. CONCLUSION: Combined analysis of tumor antigens involved in adhesion of breast cancer cells showed a negative correlation between Sialyl Lewis antigens and E-cadherin as the risk of metastasis progresses. Furthermore, there were significant differences of expression of the Sialyl Lewis antigens, E-cadherin and Cathepsin-D in primary breast cancer cells and their metastases. Evaluation of a panel of markers involved in cell adhesion could be a useful method for defining the metastatic risk in breast cancer patients.