Up-regulation of retinoblastoma protein phosphorylation in gingiva after cyclosporine A treatment: an in vivo and in vitro study.

BACKGROUND AND OBJECTIVE: Cyclosporine A can induce gingival cell proliferation; however, the precise molecular regulation of the proliferation is uncertain. Therefore, this study was carried out to examine, in vivo and in vitro, the expression of genes and proteins associated with gingival cell proliferation after treatment with cyclosporine A. MATERIAL AND METHODS: Forty Sprague Dawley rats with right maxillary posterior edentulous gingivae were assigned to a cyclosporine A group (30 mg/kg daily of cyclosporine A, administered orally) or a control group (administered mineral oil only). The animals were killed 4 wk after treatment. The edentulous gingivae were dissected out and analyzed for the expression of proliferating cell nuclear antigen (PCNA), cyclin D1, cyclin-dependent kinase 4 (CDK4) and retinoblastoma protein (Rb1) mRNA and/or protein, and phosphorylated Rb1 (pRb1), by real-time RT-PCR or immunohistochemistry. In human gingival fibroblast (HGF) cultures, the expression of PCNA, CDK4, cyclin D1 and Rb1 proteins and Rb1 phosphorylation were determined by western blotting after cyclosporine A treatment (0-10(4) ng/mL). RESULTS: Proliferating cell nuclear antigen and cyclin D1 mRNAs (Pcna and Ccnd1, respectively) were expressed more strongly in the gingivae of cyclosporine A-treated animals than in the gingivae of the controls. Immunohistochemical analyses showed that a greater number of gingival cells stained positive for cyclin D1, CDK4 and pRb1 in the cyclosporine A group than in the control group. Increased expression of cyclin D1, CDK4 and PCNA proteins was observed in HGFs after cyclosporine A treatment. The phosphorylation of Rb1 was enhanced in HGFs after treatment with cyclosporine A at concentrations of 10(2)-10(3) ng/mL. CONCLUSION: The increases in cyclin D1, PCNA and CDK4, together with the enhanced phosphorylation of Rb1, suggest that cyclosporine A promotes cell-cycle progression through the G(1)/S transition in the gingiva.

A clinical and molecular genetic study of 112 Iranian families with primary microcephaly.

BACKGROUND: Primary microcephaly (MCPH) is a genetically heterogeneous disorder showing an autosomal recessive mode of inheritance. Affected individuals present with head circumferences more than three SDs below the age- and sex-matched population mean, associated with mild to severe mental retardation. Five genes (MCPH1, CDK5RAP2, ASPM, CENPJ, STIL) and two genomic loci, MCPH2 and MCPH4, have been identified so far. METHODS AND RESULTS: In this study, we investigated all seven MCPH loci in patients with primary microcephaly from 112 Consanguineous Iranian families. In addition to a thorough clinical characterisation, karyotype analyses were performed for all patients. For Homozygosity mapping, microsatellite markers were selected for each locus and used for genotyping. Our investigation enabled us to detect homozygosity at MCPH1 (Microcephalin) in eight families, at MCPH5 (ASPM) in thirtheen families. Three families showed homozygosity at MCPH2 and five at MCPH6 (CENPJ), and two families were linked to MCPH7 (STIL). The remaining 81 families were not linked to any of the seven known loci. Subsequent sequencing revealed eight, 10 and one novel mutations in Microcephalin, ASPM and CENPJ, respectively. In some families, additional features such as short stature, seizures or congenital hearing loss were observed in the microcephalic patient, which widens the spectrum of clinical manifestations of mutations in known microcephaly genes. CONCLUSION: Our results show that the molecular basis of microcephaly is heterogeneous; thus, the Iranian population may provide a unique source for the identification of further genes underlying this disorder.

Effect of cyclosporine-A on orthodontic tooth movement in rats.

OBJECTIVE: The objective of this study is to examine the effect of cyclosporine-A (CsA) on the rate of orthodontic tooth movement in rats. SETTING AND SAMPLE POPULATION: This is a randomized controlled trial with a split-mouth design in Sprague-Dawley rats. MATERIAL AND METHODS: Eighteen rats, divided at random in two groups, were fed with 8 mg/kg CsA (experiment) or mineral oil (control) daily after initial healing of bilateral maxillary second molar removal. All rats received orthodontic coil springs (10 cN) secured to the maxillary incisors and first molars at the rights side, while no springs were placed at the left. Distances between first and third molars were measured on days 0, 3, 6, and 12. After sacrificing on day 12, the alveolar ridges of the maxillae were sectioned and blood samples were collected for serum tartrate-resistant acid phosphatase (TRAP)-5b level detection and for histology, respectively. RESULTS: Significantly larger changes in intermolar distances were found after orthodontic force application in the CsA group at days 3 and 12 when compared with the control group. The inter-radicular dental alveolus of CSA-fed rats was osteopenic. Significantly increased TRAP-5b serum level was noted in the CsA group when compared with the control group. CONCLUSIONS: We suggest that CsA enhanced the rate of orthodontic tooth movement. The osteopenia and the increased osteoclastic activity could be the underlying factors.

Cyclosporine A inhibits the expression of membrane type-I matrix metalloproteinase in gingiva.

BACKGROUND AND OBJECTIVE: Membrane type-I matrix metalloproteinase (MMP) and tissue inhibitor of metalloproteinase-2 (TIMP-2) regulate the activation of MMP-2; however, their roles in the activation of MMP-2 in gingiva during treatment with cyclosporine A are still unknown. Therefore, the expressions of membrane type-I MMP and TIMP-2, as well as MMP-2, in gingivae upon treatment with cyclosporine A were examined in vivo and in vitro. MATERIAL AND METHODS: Thirty-four rats were divided into two groups after edentulous ridges were established. The experimental group received 30 mg/kg/d of cyclosporine A and the control group received vehicle. At the end of the experimental period, the rats were killed, the gingivae were obtained and the expression of mRNA and protein of membrane type-I MMP, TIMP-2 and MMP-2 in gingiva were examined using real-time polymerase chain reaction and immunohistochemistry. In human gingival fibroblasts, the activity of MMP-2 and the expression of MMP-2, membrane type-I MMP and TIMP-2 mRNAs were examined (using zymography and reverse transcription-polymerase chain reaction, respectively) after treatment with cyclosporine A. RESULTS: In gingivae of rats, cyclosporine A significantly decreased the expression of mRNA and protein of membrane type-I MMP, but not of TIMP-2. The expression of MMP-2 mRNA was unaffected but the expression of MMP-2 protein showed a significant decrease upon treatment with cyclosporine A. In fibroblast culture medium, the presence of cyclosporine A induced a decrease in MMP-2 activity in a dose-dependent manner. The expression of MMP-2, membrane type-I MMP and TIMP-2 mRNAs in fibroblasts was not significantly affected by cyclosporine A; however, in fibroblasts the ratio of mRNA expression of membrane type-I MMP to that of TIMP-2 decreased as the cyclosporine A dose was increased. CONCLUSION: Cyclosporine A inhibits the expression of membrane type-I MMP in gingiva and it may further reduce the activation of MMP-2.

Tetracycline release from tripolyphosphate-chitosan cross-linked sponge: a preliminary in vitro study.

BACKGROUND AND OBJECTIVE: The aim of this study was to design a tripolyphosphate-chitosan cross-linked tetracycline-containing (TPP-TC) sponge that slowly releases tetracycline, for future periodontal applications. MATERIAL AND METHODS: Chitosan sponge was made by freezing and drying 2.5% chitosan solution. Tripolyphosphate-chitosan cross-linked (TPP) sponge was made by immersing the chitosan sponge in tripolyphosphate solution and air drying it. Tetracycline-containing chitosan (TC) sponge was prepared by freezing and drying a mixture of chitosan and tetracycline. TPP-TC sponge was made by immersing the TC sponge in tripolyphosphate solution. The weight, thickness and diameter of the four chitosan sponges were recorded. Their surface microstructures were inspected using scanning electron microscopy. The amount of tetracycline released from the sponges was analyzed by spectrophotometry. Antimicrobial activities of the residual sponges were tested against Staphylococcus aureus and Escherichia coli. RESULTS: The topography of the scaffolds was intact after the addition of tetracycline. However, increased surface irregularities were noted. In sponges with tripolyphosphate, intensified surface folding was observed. The weight of the sponges increased after tripolyphosphate and tetracycline were added, but their thicknesses and diameters decreased after cross-linking. Tetracycline was detected in the solution containing TPP-TC sponges until day 11. On day 7, the tetracycline released from TC sponges was less than that released from TPP-TC sponges. Bacterial growth was inhibited by sponges containing tetracycline. The inhibitory effect of the TPP-TC sponges was detectable until day 11. CONCLUSION: Our data showed that TPP-TC sponge was suitable as a slow-release device for tetracycline and that it maintained antimicrobial effects against the bacteria tested for up to 11 d.

A stereomicroscopic and immunohistochemical study of vasculature in gingiva bleeding after probing.

This study investigated the stereomicroscopic appearance of gingival sulcular surfaces and the histologic alterations of subepithelial vasculature of gingiva bleeding after probing. Sixty-four pairs of gingival biopsies from 49 patients were obtained and the results of stereomicroscopically visible vasculature were observed in half of the specimens. The number of biopsies showing that the vasculature of bleeding gingiva was significantly greater than those of non-bleeding gingiva. The immunohistochemical marker UEA-1 was used to identify endothelial cells on the vascular walls of 34 gingival biopsies from 15 patients. Following stereomicroscopic examination, tissue preparation with routine and UEA-1 stainings was performed. Volumetric and numerical analyses of vasculature in 6 divided portions of subepithelial connective tissue were measured. The results showed that significantly greater numbers of UEA-1 staining vessels were found within the apico-superficial portion of bleeding gingiva than those of non-bleeding gingiva. However, there was no specific correlation between the UEA-1 positive vessels within gingival tissue and stereomicroscopic visibility of vasculature on the surface appearance. It is suggested that increased vascularity in gingiva may be responsible for bleeding on clinical probing.

The effect of plaque retention on cyclosporine-induced gingival overgrowth in rats.

The purpose of this study was to examine the role of plaque retention on cyclosporine A (CsA)-induced gingival overgrowth. Forty-five male Sprague-Dawley rats, 15 for each of three CsA dosage conditions, were unilaterally ligated around the first mandibular molar (plaque retention). The silk ligature was left in place for 6 weeks. Contralateral first molars served as unligated controls. The daily dosage of CsA, administered by gastric feeding, was 0, 3, or 10 mg/kg body weight. Stone models from biweekly impressions of the molar sites were used to investigate development of gingival overgrowth. Rats were sacrificed at 6 weeks for histopathological and histometric examination of the molar sites. Gingival overgrowth was significantly increased in sites with higher CsA dosage, longer treatment duration, and ligation. Gingival overgrowth was enhanced in ligated sites regardless of CsA dosage. However, the odds ratio of ligated over unligated sites for gingival overgrowth increased with increasing CsA dosage. The histopathological and histometric examination revealed significantly increased gingival volume in ligated sites in CsA-treated animals. The tissue enlargement included both the epithelium and the connective tissue; however, the epithelium to connective tissue ratio remained unaltered. Within limitations of the study, we suggest that plaque retention magnifies CsA-induced gingival overgrowth; thus, dental plaque appears to be a cofactor in the development of CsA-induced gingival overgrowth.

Dose-dependent gingival overgrowth induced by cyclosporin in rats.

This study evaluated the effect of dosage on severity of cyclosporin-A (CSA) induced gingival overgrowth. Eighty (80) male Sprague-Dawley rats were randomly distributed into 4 groups. Rats in each group daily received CSA in mineral oil by gastric feeding at dosages of 0 (control), 3, 10, and 30 mg/kg, respectively, for 6 weeks. Stone models of the mandibular incisal region were obtained biweekly and were used for analysis of the gingival dimensions. Animals were sacrificed at the end of week 6 and tissue sections were processed for histopathologic evaluations. Animals were sacrificed at the end of week 6 and tissue sections were processed for histopathologic evaluation Gingival overgrowth including bucco-lingual and mesio-distal width and vertical height were significantly increased with increasing CSA dosage. Furthermore, the gingival dimensions displayed a positive linear relation to dosage and treatment duration. The histopathologic evaluation revealed a granulomatous tissue wedging the tooth-gingival interface in the 3 mg/kg group. This tissue had reached exuberant size in the 10 and 30 mg/kg groups. In summary, the analysis of gingival dimensions the histopathologic evaluation shows a dose-dependent effect on the severity of CSA-induced gingival overgrowth.

Nifedipine-induced gingival overgrowth in rats: brief review and experimental study.

The first case report of gingival overgrowth induced by nifedipine (NIF), a calcium-beta blocker, was in 1984. However, the association between gingival alterations and the drug therapy of sodium diphenyl hydantoinate was initially described in 1939. The purpose of the experimental study was to examine the effect of NIF on gingival morphology in an animal model. Forty-five male Sprague-Dawley rats were randomly divided into 3 groups. Animals in each group daily received NIF in dimethyl sulfoxide by gastric feeding at a dosage of 0 (control), 30, or 50 mg/kg body weight for 9 weeks. Gingival gross morphology was assessed tri-weekly from stone models obtained from the mandibular incisal region. Animals were sacrificed at the end of study and tissue blocks were processed for histopathologic and histometric evaluation. Histometric analysis was performed at 5 selected tissue levels. Macro- and microscopic significantly increased gingival dimensions were demonstrated in NIF-treated animals compared to control. Although a fibrovascular tissue was observed in the tooth-gingiva interface for both NIF-treated and control animals, it was thicker and appeared earlier in NIF-treated animals. The results of the present study suggest that gingival overgrowth can be induced by NIF in rats and that the gingival overgrowth appears dose dependent.

Effects of cyclosporin A on alveolar bone: an experimental study in the rat.

BACKGROUND: There have been several investigations on the role of cyclosporin A (CSA) in gingival hyperplasia in both animals and humans. However, less attention has been given to the drug's effect on alveolar bone. This study used light microscopy to histologically and histometrically evaluate the effects of CSA on alveolar bone in the rat. METHODS: Sixty, 6-week-old Sprague-Dawley rats were separated into test and control groups. Animals in the test group received CSA in mineral oil (30 mg/kg body weight) daily by gastric feeding over the 6-week study. Control animals received only mineral oil. Ten animals from each group were sacrificed at weeks 2, 4, and 6. After histologic processing, the labial crest of the alveolar bone in the anterior mandible was evaluated by light microscopy. RESULTS: A distinct pattern of increased osteoclasia and reduced bone formation was observed in the CSA-exposed animals compared to the controls. Increased osteoclasia was observed in periodontal sites and decreased bone formation was observed in symphyseal sites. CONCLUSIONS: The results suggest that CSA has distinct effects on alveolar bone.

Gingival overgrowth and dental alveolar alterations: possible mechanisms of cyclosporin-induced tooth migration. An experimental study in the rat.

The inter-incisal distance and dimension of the interdental papilla between the mandibular incisors were examined in cyclosporin A (CSA) fed rats over 6 weeks. Rats in the test group received CSA daily in mineral oil by gastric feeding (30 mg/kg body weight); the control group received mineral oil only. The inter-incisal distance and gingival dimensions, including bucco-lingual width and vertical height, were assessed biweekly from alginate impressed stone models. Animals were sacrificed at the end of the study and tissue sections were obtained from the anterior region of the mandible for histopathological evaluation. Both the inter-incisal distance and the dimension of the interdental papilla were significantly greater in CSA-exposed animals compared to control. The significant alteration appeared earlier in the papillary dimensions than that in the interdental distance. Particular histopathological alterations of the soft and hard tissues of the periodontium were observed in CSA-exposed animals. Within limitations of the study, we suggest that the CSA-induced gingival overgrowth may offer an active force contributing to observed tooth movement, however, remarkable alveolar remodeling should be considered as an undetermined factor for the movement.

Cyclosporin-induced gingival overgrowth at the newly formed edentulous ridge in rats: a morphological and histometric evaluation.

BACKGROUND: Since cyclosporin A (CsA)-induced overgrowth seldom occurs at sites distant from teeth, the periodontal ligament has been considered significant. The aim of this study was to examine overgrowth occurrence at the edentulous ridge--the sites without the ligament--after CsA therapy in rats. METHODS: After extracting all right maxillary molars, 16 Sprague-Dawley rats underwent a 2-week healing period. The animals were separated into CsA and control groups. CsA rats received 15 mg/kg of CsA by gastric feeding for 4 weeks, while the control group received only mineral oil. At the end of study, all animals were sacrificed and stone models were immediately obtained by rubber-based impressions. The edentulous ridge morphology, including the bucco-lingual width and the vertical height, was measured on the models. For histometry, 10 sections were selected from the edentulous ridge of each animal after undecalcified tissue preparation. The soft tissue areas of the edentulous ridge and the trabecular bone morphology of the dental alveolus were measured. RESULTS: CsA therapy produced a significant increase of the ridge width and height, measured from the stone models, when compared to the control group. Under histometry, CsA resulted in a significant increase of the epithelium, connective tissue, and total soft tissue areas. The measured trabecular bone volume was affected by both examining factors: the drug therapy and the location of the dental alveolus. CsA therapy produced a significant loss of bone volume but a significant increase of the bone-specific surface area. Although the mean osteoid volume was similar between CsA and control groups, a significant decrease of the fractional formation surface in the CsA group was revealed. CONCLUSIONS: An enlarged edentulous ridge and an altered dental alveolar bone morphology were observed in CsA-treated animals at the end of the study; therefore, we suggest that CsA may induce not only a soft tissue overgrowth but also an alveolar bone alteration at the edentulous ridge. The hypothesis that tooth or periodontal ligament is an essential component for the overgrowth development is questioned.

Effects of cyclosporin A on the mandibular condylar cartilage in rats.

Twenty, 5-week-old, male, Sprague-Dawley rats were divided into a control and a cyclosporin A (CSA) group for evaluating effects of the drug on condylar cartilage. Animals in the treatment group daily received CSA (15 mg/kg body wt) in mineral oil by gastric feeding over a 4-week observation interval. Control animals received mineral oil only. Five animals from each group were killed at weeks 2 and 4 of study. After histological processing, five tissue sections from the mid-region of the condyle were selected and examined. Three compositional zones (articular fibrous, proliferative, and hypertrophic) of the superior, posteriosuperior and posterior regions of the condylar cartilage were evaluated by light microscopy. At week 2, total condylar cartilage thickness was similar in the CSA and control groups, but the thickness of each zone was altered in CSA-treated animals, including a decrease of the fibrous and proliferative zones and an increase in hypertrophic zone compared to control (P<0.05). At week 4, CSA-treated animals exhibited overall decreased cartilage thickness, including decreased thickness of each zone compared to control (P<0.05). The results suggest that CSA has an inhibitory effect on the maturation of the mandibular condyle in rats.

Viral load of high-risk human papillomavirus in cervical squamous intraepithelial lesions.

OBJECTIVES: This case-control study was conducted to investigate the role of viral load of high-risk human papillomaviruses (HPVs) in the development of cervical squamous intraepithelial lesions (SILs) and invasive cancers. METHODS: A total of 30 female cases who had histological evidence of low-grade SIL (n=10) or high-grade SIL and above (n=20) were identified as the case group at the Tri-Service General Hospital, Taipei between September 1998 and March 1999. In addition, 80 female controls who had normal cervical cytology were enrolled and individually matched on age (+/-5 years) and date of recruitment to each case. Cervical swabs collected from study subjects were tested for the positivity and viral load of high-risk HPVs by Hybrid Capture II assay. Additionally, subjects completed a risk factor questionnaire. RESULTS: Among sex behavioral factors studied, younger age at first intercourse was associated with a significantly elevated risk of cervical SIL and invasive cancers. With respect to HPV infection, high-risk HPV DNA was present in 70% (21/30) of case and 21% (17/80) of control subjects, resulting in an odds ratio (OR) of 6.6 [95% confidence interval (C.I.)=2.6-17.0]. Moreover, women who had a high viral load were at significantly greater risk for cervical SIL and invasive cancers than those who were infected with a low viral load (OR=18.0, 95% C.I.=3.0-108.5). CONCLUSIONS: Among the variables tested, infection with a high viral load of high-risk HPVs is the strongest determinant for cervical SIL and cervical cancers in Taiwan.

Cytology of fine needle aspirates of primary extragonadal germ cell tumors.

OBJECTIVE: To describe the characteristic cytologic features of fine needle aspirates (FNAs) of primary extragonadal germ cell tumors (PEGCTs). STUDY DESIGN: Thirteen patients with PEGCTs, including 2 seminomas, 2 mixed germ cell tumors, 3 immature teratomas, 1 choriocarcinoma and 5 yolk sac tumors (YSTs) were studied. The final diagnosis of PEGCT in all cases was established by histologic examination of the tumor tissues. Fine needle aspiration was done on either the primary tumor or metastatic foci. The aspirates were stained with one of the Romanovsky stains and Papanicolaou stain. RESULTS: Each type of PEGCT has its own morphologic characteristics. In seminoma, the tumor cells are large and noncohesive, with one to several distinct nucleoli; some lymphocytes are also present. YSTs show many pleomorphic cells with vacuoles in the cytoplasm and nuclei; tumor cells frequently aggregate in a microglandular or papillary pattern. Choriocarcinoma consists of syncytiotrophoblasts and cytotrophoblasts. The former are very large cells with eosinophilic cytoplasm, one to several nuclei and distinct nucleoli; the latter are medium-sized cells with vacuolated, basophilic cytoplasm and eccentric nuclei. Immature teratomas are composed of a mixture of cell types, including elongated epithelioid cells, mesenchymal cells and many large, naked, amorphous nuclei with a homogeneous chromatin pattern. Diagnosis of mixed germ cell tumor is difficult but can be made if two or more subtypes of tumor cells are observed in the FNA. CONCLUSION: Cytologic examination of FNAs of primary or metastatic lesions of PEGCTs, stained either with Romanovsky or Papanicolaou stain, is of diagnostic value for such diseases. The use of immunochemistry can help to confirm the cytologic impression.

Diagnosis of primary cardiac lymphoma. Report of a case with cytologic examination of pericardial fluid and imprints of transvenously biopsied intracardiac tissue.

BACKGROUND: Primary cardiac lymphoma (PCL) is a treatable disease when appropriately diagnosed. Therefore, a prompt, safe method with high diagnostic accuracy is prerequisite to successful therapy for PCL. CASE: A 57-year-old male presented with exertional dyspnea and atrial fibrillations. A pericardial effusion (PE) and several tumor masses occupying both atria were found. Cytologic examinations of PE and of imprints of the tissues obtained by transvenous biopsy of the cardiac tumors revealed numerous small, round tumor cells and lymphoglandular bodies, suggestive of malignant lymphoma. This cytologic impression was confirmed by immunocytochemical studies on the same cytologic material. Histologic studies reaffirmed the diagnosis of B-cell lymphoma. The patient received eight courses of chemotherapy, with complete remission of the illness. CONCLUSION: Cardiac lymphoma can be quickly and safely diagnosed by cytologic examination of PE or transvenously biopsied cardiac tissue, with confirmation by immunocytochemical studies. Exploratory thoracotomy for biopsy can be avoided.

Inflammatory pseudotumor of the liver with occlusive endophlebitis: report of a case.

Inflammatory pseudotumor (IPT) of the liver is an extremely rare benign tumor. Only 12 cases have so far been reported in the English literature. The etiology and pathogenesis of IPT remain obscure. We herein present an additional case of IPT of the liver with occlusive endophlebitis in a 36-year-old female. Immunohistochemical studies demonstrated the polyclonal nature of the plasma cells in the tumor, including IgA, IgM, IgG, kappa and lambda. The inflammatory pseudotumor should be kept in mind in the differential diagnosis of hepatic space-occupying malignant lesions.

Cyclosporin A-induced gingival overgrowth in rats: macroscopic and microscopic observations.

Because the existence of cyclosporin A-induced gingival overgrowth in animals has not been well established, a pilot study was undertaken to determine whether such gingival overgrowth could be observed in Sprague-Dawley rats. Thirty-six male rats, age 6 weeks, were randomly divided into two groups. The test group was given 30 mg/kg of body weight of cyclosporin A daily by gastric feeding. The control group was given mineral oil instead. Stone casts of the gingiva in the mandibular incisor region were made biweekly for 6 weeks. The three-dimensional growth pattern-the buccolingual width, the mesiodistal width, and the vertical height of the gingiva-was analyzed on the casts. The animals were killed after the last impression was taken, and tissue sections were made. Stereomicroscopy revealed marked overgrowth of the gingiva in the test rats. All three-dimensional measurements on the stone casts were greater in the test group, beginning 2 weeks after cyclosporin A was given. Histologically the overgrowth of the epithelia and connective tissue was easily confirmed in the buccal and lingual gingiva of the test rats.

Clinical significance of tumor suppressor PTEN in colorectal carcinoma.

BACKGROUND: It has been demonstrated that the deletion, mutation, hypermethylation and subcellular location of the tumor suppressor phosphatase and tensin homologue (PTEN) are closely correlated with carcinogenesis, progression and prognosis of malignancy. Both mutation and the microsatellite instability of the PTEN gene influence regulation of the PI3K/Akt signaling pathway. This study investigated whether loss of nuclear PTEN is correlated with chemosensitivity, clinicopathological parameters and survival. METHODS: Intracellular levels of PTEN of multiple cell lines of colorectal carcinoma (CRC) were evaluated by Western blotting and immunocytochemistry. The chemosensitivity of cell lines with various expression levels of PTEN was evaluated using 5-flurouracil (5-FU), oxaliplatin and irinotecan (CPT), and clinical significance was evaluated by immunohistochemical analysis of 133 CRC specimens. RESULTS: Colon cancer cell lines HT-29, LoVo and SW480 differed in expression of PTEN, with high, moderate and low levels, respectively. HT-29 and LoVo PTEN expression was suppressed by a low concentration of 5-FU and oxaliplatin; however, SW480 was insensitive to these chemotherapeutic agents. Nuclear PTEN was overexpressed in most (>80%) normal colon mucosa samples, but the incidence significantly decreased (89.2% → 53.4%) in the CRC group. PTEN in the nucleus was negatively correlated with tumor size and vascular invasion in CRC, and CRC patients with negative PTEN expression in the nucleus exhibited poor survival. CONCLUSION: Cell lines with a high expression of PTEN are sensitive to chemotherapy with 5-FU and oxaliplatin. Nuclear PTEN expression gradually decreases after malignant transformation, and loss of PTEN expression in the nucleus is associated with tumor progression and poor clinical outcome in CRC.

Nrf-2 regulates cyclosporine-stimulated HO-1 expression in gingiva.

Cyclosporine-A (CsA) stimulates heme oxygenase-1 (HO-1) expression in the gingiva, but the regulation and the role of HO-1 in gingival overgrowth are not well-understood. HO-1 is regulated by several transcription factors, such as nuclear factor-κB (NF-κB) and nuclear factor erythroid-2-related factor 2 (Nrf-2). The aim of this study was to examine the role of Nrf-2 in the regulation of CsA-stimulated HO-1 expression in human gingival fibroblasts. Nrf-2 siRNA (siNrf-2), NF-κB, kinase inhibitors, and sulforaphane (SFN) were used to examine the nuclear translocation of Nrf-2 and expression of HO-1 and transforming growth factor-ß1 (TGF-ß1) in cells. Treatment with siNrf-2, but not with an NF-κB inhibitor, reduced CsA-stimulated HO-1 mRNA expression. ERK inhibition significantly decreased CsA-stimulated Nrf-2 nuclear translocation and HO-1 mRNA expression. Pre-treatment with SFN showed that HO-1 plays a role in attenuating CsA-mediated TGF-ß1 expressions. These findings suggest that CsA-stimulated HO-1 expression is mediated through the activation of ERK, and that Nrf-2 plays a protective role against CsA-induced gingival fibrosis by modulating collagen turnover-related genes.