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.

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.

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.

Surgical treatment of gastrointestinal stromal tumor in the esophagus: report of three cases.

BACKGROUND/AIMS: Gastrointestinal stromal tumors (GISTs) are rare mesenchymal tumors of the alimentary tract. The term GIST was introduced in 2004 and understanding of the tumor's cellular origin, classification, diagnostic markers, and prognostic parameters has evolved dramatically over the last two decades. Hirota et al. proposed that GISTs originate from interstitial cells of Cajal - regulators of gut peristalsis that normally express CD 117 - which is the product of the c-kit proto-oncogene that encodes a tyrosine kinase receptor that regulates cellular proliferation in GISTs. In the esophagus, squamous cell carcinoma and adenocarcinoma are common malignant tumors and leiomyoma is the most frequent mesenchymal neoplasm. Esophageal GISTs, however, have been reported less frequently. METHODS: We report three cases of esophageal GISTs in patients who underwent surgical intervention in our institution. The patients suffered from dysphagia, without specific findings on initial physical examination. Submucosal tumors were suspected after the patients underwent barium swallow and endoscopic studies. In addition, positron emission tomography was used to study a submucosal tumor in one patient. RESULTS: The pathological diagnosis was confirmed in all cases by microscopic examination with hematoxylin and eosin stain and positive immunoreaction for c-kit. Two of them were low risk and the third one was high risk in character, according to the consensus approach and depending on the size and mitotic index of the tumor. CONCLUSION: The patients had uneventful postoperative recoveries and were followed up regularly at 3-month intervals.

Expression of fascin in oral and oropharyngeal squamous cell carcinomas has prognostic significance - a tissue microarray study of 129 cases.

AIMS: To elucidate the role of fascin in oral and oropharyngeal squamous cell carcinoma (OSCC) by correlation with clinical parameters. METHODS AND RESULTS: Paraffin sections using tissue microarrays of 129 patients with OSCC were investigated immunohistochemically. Fascin protein was overexpressed in OSCC cells compared with their non-neoplastic epithelial counterparts. For evaluating the intensity of fascin, 39 (30.2%) were classified as weakly immunoreactive, 76 (58.9%) as moderate reactive and 14 (10.9%) as intensely reactive. For evaluating the distribution of fascin, 64 (49.6%) were classified as < 55% and 65 (50.4%) were classified as >/= 55%. Fascin protein expression was correlated with size or extent of the tumour (P < 0.001), positive lymph node metastasis (P < 0.001), distant metastasis (P = 0.014) and clinical staging (P < 0.001). The immunoreactivity scores of fascin in OSCC were variable but showed significant correlation with histological grade, clinical TNM system and stage. CONCLUSION: Expression of fascin protein may play an important role in progression of OSCC. Overexpression of fascin contributes to a more aggressive clinical course and suggests the potential of fascin as a new molecular target for therapeutic intervention.

Differential viral loads of human papillomavirus 16 and 58 infections in the spectrum of cervical carcinogenesis.

Human papillomavirus (HPV) load was reported to be related to the severity of cervical neoplasia but with controversy. The viral load-disease severity relationship was showed in HPV 16, but no study was made in HPV 58, the second most prevalent HPV in cervical cancer in East Asia. We studied cervical HPV loads in HPV 16- and HPV 58-infected cases of normal, low-grade squamous intraepithelial lesions (LSIL), high-grade squamous intraepithelial lesions (HSIL), and invasive cervical cancer (CC) by using quantitative polymerase chain reaction (Q-PCR) with type-specific primers in defined cell number. With the exception of HPV 16 infection in normal, viral loads varied greatly in each disease regardless of genotypes. The load of HPV 16 differed significantly among disease severities, with a dramatic increase from normal (1.14 +/- 2.25 copies/cell) to LSIL, HSIL, and CC (1599 +/- 2301, 7489 +/- 24,087 and 1878 +/- 2979 copies/cell, respectively) (P < 0.01). No significant difference was noted among different HPV 58 infections, with loads in normal, LSIL, HSIL, and CC of 503 +/- 641, 7951 +/- 27,557, 353 +/- 744, and 1139 +/- 2895 copies/cell, respectively. In comparison with HPV 16, HPV 58 subclinical infection confers a significant higher load (P < 0.01). Different HPV types behave differentially in the spectrum of cervical carcinogenesis. Unlike HPV 16, the infection load of HPV 58 does not correlate to the clinical severity. The wide variation of HPV loads among different HPV types and among squamous intraepithelial lesions and CC makes the viral load test unrealistic in differentiating different severities of cervical neoplasia.

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.

Examination of pericardial effusions by cytology and immunocytochemistry.

BACKGROUND: Cytologic examination of pericardial effusions (PE) has been uncommonly reported in the literature. Fewer reports have studied the role of immunocytochemistry in diagnosis of PE. We described our experience, according to cytologic examination and immunocyto-chemical staining of PE. METHODS: Over a four-year period, 50 PE specimens from 36 patients were examined cytologically by Wright-Giemsa stain and/or Papanicolaou stain, at Tri-Service General Hospital. Immunocytochemical stainings were further performed to ensure the cell nature. RESULTS: Eighteen of these patients had benign etiologies; the others suffered from malignant diseases. Sixteen cases, including eight benign and eight malignant, received pericardial biopsy for comparison. Commonly encountered cells in PE with any kind of etiology included neutrophils, lymphocytes macrophages and mesothelial cells. Cancer cells were additionally seen in malignant PE, but not in all cases with cancers. Furthermore, reactive mesothelial cells and macrophages were sometimes not easily distinguished from cancer cells, and reactive lymphocytes frequently mimicked hematopoietic malignancies. Further, the nature of small round tumor cells can hardly be ensured by morphologic examination only. Immunocytochemical studies were successfully used to solve these dilemmas in some suspicious cases. CONCLUSIONS: This study showed that cytology together with clinical information and immunocytochemistry can achieve excellent sensitivity and specificity in identification of malignant PE. The use of immunocytochemistry can even ensure the nature of certain cancers such as small cell carcinoma and B cell lymphoma.

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.

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.

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.