Uterine angioleiomyoma with disseminated intravascular coagulation: a case report.

BACKGROUND: Uterine angioleiomyoma is benign tumor that composed of smooth muscle cells and thick-walled vessels. It is a very rare condition reported to present as lower abdominal mass, accompanied by dysmenorrhea and hypermenorrhea. However, its clinical presentation is not known. CASE PRESENTATION: We report the case of a 44-year-old Japanese woman who developed severe anemia with disseminated intravascular coagulation without obvious external bleeding. The patient had a huge abdominal mass of over 20 cm in size, which was thought to be a uterine tumor. She received daily blood transfusions and her condition improved rapidly after she underwent hysterectomy. Pathological examination of the tumor revealed spindle-shaped cells with little atypia and mitosis, and numerous large vessels with smooth muscle and thrombus in the vessels. CONCLUSIONS: Uterine angioleiomyoma was identified as the cause of the coagulation abnormality. CCND2 and AR gene amplification was detected in the tumor. Uterine tumors that present with coagulopathy despite a clinical course suggestive of benign disease should undergo differential diagnosis for uterine angioleiomyoma.

Associations of streptococci and fungi amounts in the oral cavity with nutritional and oral health status in institutionalized elders: a cross sectional study.

BACKGROUND: Disruption of the indigenous microbiota is likely related to frailty caused by undernutrition. However, the relationship between undernutrition and the oral microbiota, especially normal bacteria, is not obvious. The aim of this study was to elucidate the associations of nutritional and oral health conditions with prevalence of bacteria and fungi in the oral cavity of older individuals. METHODS: Forty-one institutionalized older individuals with an average age ± standard deviation of 84.6 ± 8.3 years were enrolled as participants. Body mass index (BMI) and oral health assessment tool (OHAT) scores were used to represent nutritional and oral health status. Amounts of total bacteria, streptococci, and fungi in oral specimens collected from the tongue dorsum were determined by quantitative polymerase chain reaction (PCR) assay results. This study followed the STROBE statement for reports of observational studies. RESULTS: There was a significant correlation between BMI and streptococcal amount (ρ = 0.526, p < 0.001). The undernutrition group (BMI < 20) showed a significantly lower average number of oral streptococci (p = 0.003). In logistic regression models, streptococcal amount was a significant variable accounting for "not undernutrition" [odds ratio 5.68, 95% confidential interval (CI) 1.64-19.7 (p = 0.06)]. On the other hand, participants with a poor oral health condition (OHAT ≥ 5) harbored significantly higher levels of fungi (p = 0.028). CONCLUSION: Oral streptococci were found to be associated with systemic nutritional condition and oral fungi with oral health condition. Thus, in order to understand the relationship of frailty with the oral microbiota in older individuals, it is necessary to examine oral indigenous bacteria as well as etiological microorganisms.

Adenovirus vector expressing keratinocyte growth factor using CAG promoter impairs pulmonary function of mice with elastase-induced emphysema.

Pulmonary emphysema impairs quality of life and increases mortality. It has previously been shown that administration of adenovirus vector expressing murine keratinocyte growth factor (KGF) before elastase instillation prevents pulmonary emphysema in mice. We therefore hypothesized that therapeutic administration of KGF would restore damage to lungs caused by elastase instillation and thus improve pulmonary function in an animal model. KGF expressing adenovirus vector, which prevented bleomycin-induced pulmonary fibrosis in a previous study, was constructed. Adenovirus vector (1.0 × 109 plaque-forming units) was administered intratracheally one week after administration of elastase into mouse lungs. One week after administration of KGF-vector, exercise tolerance testing and blood gas analysis were performed, after which the lungs were removed under deep anesthesia. KGF-positive pneumocytes were more numerous, surfactant protein secretion in the airspace greater and mean linear intercept of lungs shorter in animals that had received KGF than in control animals. Unexpectedly, however, arterial blood oxygenation was worse in the KGF group and maximum running speed, an indicator of exercise capacity, had not improved after KGF in mice with elastase-induced emphysema, indicating that KGF-expressing adenovirus vector impaired pulmonary function in these mice. Notably, vector lacking KGF-expression unit did not induce such impairment, implying that the KGF expression unit itself may cause the damage to alveolar cells. Possible involvement of the CAG promoter used for KGF expression in impairing pulmonary function is discussed.

Class III/IV POU transcription factors expressed in small cell lung cancer cells are involved in proneural/neuroendocrine differentiation.

One-third of lung malignancies demonstrate a proneural/neuroendocrine phenotype or type of differentiation. However, it has not been clearly elucidated how proneural/neuroendocrine differentiation is controlled in lung cancers. We recently demonstrated that the POU3F2 gene plays a significant role in proneural/neuroendocrine differentiation of lung cancers. Because class III POU genes (POU3F1, POU3F2, POU3F3, and POU3F4) and class IV POU genes (POU4F1, POU4F2, and POU4F3) share similar properties in neural development, we analyzed the association between class III/IV POU genes and a proneural/neuroendocrine phenotype in lung cancers using seven small cell lung cancer (SCLC) cell lines and twelve non-SCLC (NSCLC) cell lines. Class III/IV POU gene expression was generally restricted to SCLC cells. However, the forced expression of class III/IV POU genes in the NSCLC cell lines induced the expression of neuroendocrine-specific markers (neural call adhesion molecule 1, synaptophysin, and chromogranin A) and proneural transcription factors (achaete-scute homolog-like 1, NeuroD1, and thyroid transcription factor 1) in various degrees. Furthermore, each class III/IV POU gene induced other class III/IV POU genes, suggesting the mutual induction of class III/IV POU genes. These findings suggest that the expression of class III/IV POU genes is important for the proneural/neuroendocrine differentiation of lung cancer cells.

Expression of developing neural transcription factors in lung carcinoid tumors.

In lung tumors, the association between carcinoids and high-grade neuroendocrine tumors (HGNETs) is controversial. To understand the phenotypic similarities/differences between lung carcinoids and HGNETs, we comparatively investigated the expression of three kinds of developing neural transcription factors (DNTFs: BRN2, TTF1 and ASCL1) and multiple endocrine neoplasia type 1 (MEN1) as well as RB1 and P53 using 18 carcinoids and 16 HGNETs. The DNTFs were expressed in 10 of the 18 carcinoids and in all the HGNETs, while normal neuroendocrine cells, which are considered the major cell origin of lung carcinoids and small cell carcinomas, did not express DNTFs. Both the DNTF(-) and DNTF(+) carcinoids contained typical and atypical carcinoids. All the DNTF(-) carcinoids examined were formed in the bronchial wall. All the MEN1(-) carcinoids examined were classified into the DNTF(-) carcinoids, while all the HGNETs expressed MEN1. This finding suggests that DNTF(-) MEN1(-) carcinoids are unlikely to be precursors of HGNETs. Although the status of RB1 and P53 between carcinoids and HGNETs were apparently different, the DNTF(+) carcinoids of two male patients and one female patient revealed morphologies resembling HGNET cells and relatively high Ki67 indices. Further investigation of DNTF expression in carcinoids might provide important clues to understand the association between carcinoids and HGNETs.

Neural lineage-specific homeoprotein BRN2 is directly involved in TTF1 expression in small-cell lung cancer.

Thyroid transcription factor 1 (TTF1) plays crucial roles in thyroid, lung, and developing brain morphogenesis. Because TTF1-expressing neoplasms are generated from organs and tissues that normally express TTF1, such as the thyroid follicular epithelium and peripheral lung airway epithelium, TTF1 is widely used as a cell lineage-specific and diagnostic marker for thyroid carcinomas and for lung adenocarcinomas with terminal respiratory unit (TRU) differentiation. However, among lung neuroendocrine tumors, small-cell carcinomas (small-cell lung cancers (SCLCs)), most of which are generated from the central airway, also frequently express TTF1 at high levels. To clarify how SCLCs express TTF1, we investigated the molecular mechanisms of its expression using cultivated lung cancer cells and focusing upon neural cell-specific transcription factors. Both SCLC cells and lung adenocarcinoma cells predominantly expressed isoform 2 of TTF1, and TTF1 promoter assays in SCLC cells revealed that the crucial region for activation of the promoter, which is adjacent to the transcription start site of TTF1 isoform 2, has potent FOX-, LHX-, and BRN2-binding sites. Transfection experiments using expression vectors for FOXA1, FOXA2, LHX2, LHX6, and BRN2 showed that BRN2 substantially upregulated TTF1 expression, whereas FOXA1/2 weakly upregulated TTF1 expression. BRN2 and FOXA1/2 binding to the TTF1 promoter was confirmed through chromatin immunoprecipitation experiments, and TTF1 expression in SCLC cells was considerably downregulated after BRN2 knockdown. Furthermore, the TTF1 promoter in SCLC cells was scarcely methylated, and immunohistochemical examinations using a series of primary lung tumors indicated that TTF1 and BRN2 were coexpressed only in SCLC cells. These findings suggest that TTF1 expression in SCLC is a cell lineage-specific phenomenon that involves the developing neural cell-specific homeoprotein BRN2.

POU domain transcription factor BRN2 is crucial for expression of ASCL1, ND1 and neuroendocrine marker molecules and cell growth in small cell lung cancer.

BRN2 is a developmental neural cell-specific POU domain transcription factor and is crucial for cell lineage determination. We investigated the importance of BRN2 in the expression of the lineage-specific transcription factors (achaete-scute homolog-like 1 (ASCL1) and NeuroD1 (ND1)) and neural/neuroendocrine marker molecules (neural cell adhesion molecule 1 (NCAM1), synaptophysin (SYP) and chromogranin A (CHGA)) in small cell lung cancer (SCLC) using cultured lung cancer cells. All examined SCLC cell lines expressed BRN2, as well as ASCL1, ND1, NCAM1, SYP and CHGA. The expression levels of ASCL1, ND1, NCAM1, SYP and CHGA considerably decreased when BRN2 was knocked down in SCLC cells, and the addition of a BRN2 transgene into non-SCLC (NSCLC) cells induced the expression of ASCL1, ND1, NCAM1, SYP and CHGA. However, the BRN2 gene was not activated by the forced expression of ASCL1 or ND1 in NSCLC cells. The knockdown of BRN2 caused significant growth retardation with decrease of S to G2 phase population and mitotic cell rates and unaltered Ki-67-labeled or apoptotic cell rates in SCLC cells, indicating increase of G1 phase population. These findings suggest that BRN2 is a higher level regulator than ASCL1 and ND1 and BRN2 might be involved in aggressiveness of SCLC.

Diagnostic Utility of Hysteroscopic Biopsy in Cases of Suspected Lobular Endocervical Glandular Hyperplasia and Comparison with Cervical Conization.

BACKGROUND: Cervical cystic lesions encompass a range of benign and malignant pathologies. Magnetic resonance imaging or cytology alone cannot provide a definitive diagnosis, and conventional practice involves performing a cervical biopsy by conization to confirm the histology in cases exhibiting potential signs of lobular endocervical glandular hyperplasia (LEGH) or malignancy. However, as postoperative complications resulting from conization can impact future fertility and pregnancy, alternative diagnostic methods are needed for reproductive-age patients. This study aimed to establish the efficacy of a hysteroscopic biopsy for diagnosing cervical cystic lesions and compare it with conization. METHODS: Thirteen patients with cervical cystic lesions suspected of LEGH or malignancy underwent a hysteroscopic biopsy, while 23 underwent conization. Patient background information, preoperative evaluation, histology, and postoperative outcomes were collected and compared retrospectively. RESULTS: No significant differences were found between the hysteroscopy and conization groups in terms of mean patient age (45 vs. 48 years), operating time (23 vs. 35 min), blood loss (small amount vs. 43 mL), and postoperative hospitalization (1.1 vs. 1.6 days). CONCLUSION: A hysteroscopic biopsy allows for targeted resection of the cervix while maintaining diagnostic accuracy. It may serve as an efficient method for diagnosing cervical cystic lesions.

Differences of molecular expression mechanisms among neural cell adhesion molecule 1, synaptophysin, and chromogranin A in lung cancer cells.

Neural cell adhesion molecule 1 (NCAM1), synaptophysin (SYPT), and chromogranin A (CGA) are immunohistochemical markers for diagnosing lung neuroendocrine tumors (LNETs). However, the precise expression mechanisms have not been studied in enough detail. The purpose of the present study is to define the molecular mechanisms of NCAM1, SYPT, and CGA gene expressions, using cultivated lung cancer cells and focusing upon NeuroD1 (ND1), achaete-scute homolog-like 1 (ASCL1), and known transcription factors, repressor element 1 (RE1)-silencing transcription factor (REST) and c-AMP responsive element-binding protein (CREB). Promoter assays, chromatin immunoprecipitation, and transfection experiments revealed that ND1 activated NCAM1, that ASCL1 weakly upregulated SYPT expression, and that CGA expression was not regulated by ND1 or ASCL1. REST expression was restricted in non-small cell lung cancer (NSCLC) cells, and knockdown of REST could cause as much SYPT expression as in SCLC cells and weak CGA expression in NSCLC cells. However, CGA gene upregulation via CREB activation was not found in REST-lacking NSCLC cells, indicating the requirement of some additional mechanism for sufficient expression. These results suggest that NCAM1, SYPT and CGA expressions are differently regulated by neuroendocrine phenotype-specific transcription factors and provide a reason why NCAM1 and SYPT are frequently expressed in LNETs, irrespective of malignancy grade.

Tubulovillous adenoma with high-grade dysplasia of the vulva harboring high tumor mutational burden and cancer-associated mutations: a case report.

BACKGROUND: Vulvar cancer is a rare disease, accounting for approximately 5% of gynecological malignancies. Primary adenocarcinoma of intestinal-type of the vulva or its precancerous lesion is extremely rare, and details regarding its origin, evolution and related genetic mutations are unknown. Treatment options for this cancer have not been defined. CASE PRESENTATION: A 63-year-old Japanese woman came to the hospital because she was aware of a vulvar mass. There was a 1 cm mass on the dorsal side of the vulva, just outside the remains of the hymen. Biopsy revealed suspected adenocarcinoma, and wide local excision was performed. From histopathology and immunohistochemistry, the specimen was diagnosed as tubulovillous adenoma with high-grade dysplasia of the vulva. No other primary lesions were found, and the vulva was considered the primary site. A gene panel test (FoundationOneCDx assay) showed a high tumor mutational burden and mutations in TP53, KEL, RB1, RNF43, PTEN, GNAS, and PIK3CA. CONCLUSIONS: The current case of tubulovillous adenoma with high-grade dysplasia of the vulva had a variety of cancer-associated mutations, despite being a precancerous lesion. In cases of intestinal-type neoplasms of the vulva, it may be helpful to check tumor mutational burden and gene mutations for treatment selection.

Keratinocyte growth factor gene transduction ameliorates pulmonary fibrosis induced by bleomycin in mice.

Pulmonary fibrosis has high rates of mortality and morbidity, but there is no established therapy at present. We demonstrate here that bleomycin-induced pulmonary fibrosis in mice is ameliorated by intratracheal administration of keratinocyte growth factor (KGF)-expressing adenovirus vector. Progressive pulmonary fibrosis was created by continuous subcutaneous administration of 120 mg/kg of bleomycin subcutaneously using an osmotic pump twice from Day 1 to 7 and Day 29 to 35. The mice initially exhibited subpleural fibrosis and then exhibited advanced fibrosis in the parenchyma of the lungs. These histopathological changes were accompanied by reduced lung compliance (0.041 ± 0.011 versus 0.097 ± 0.004; P < 0.001), reduced messenger expression of surfactant proteins, and reduced KGF messenger expression in the lungs at 4 weeks compared with naive group. Intratracheal instillation of Ad-KGF at 1 week after the first administration of bleomycin increased KGF mRNA expression in the lungs compared with the fibrosis-induced mice that received saline alone. The phenotype was associated with alveolar epithelial cell proliferation, increased pulmonary compliance (0.062 ± 0.005 versus 0.041 ± 0.011; P = 0.023), and decreased mortality (survival rate on Day 56: 68.8% versus 0%; P = 0.002), compared with mice receiving only the saline vehicle. These observations suggest the therapeutic utility of a KGF-expressing adenoviral vector for pulmonary fibrosis.

Early growth response-1 induces and enhances vascular endothelial growth factor-A expression in lung cancer cells.

Vascular endothelial growth factor-A (VEGF-A) is crucial for angiogenesis, vascular permeability, and metastasis during tumor development. We demonstrate here that early growth response-1 (EGR-1), which is induced by the extracellular signal-regulated kinase (ERK) pathway activation, activates VEGF-A in lung cancer cells. Increased EGR-1 expression was found in adenocarcinoma cells carrying mutant K-RAS or EGFR genes. Hypoxic culture, siRNA experiment, luciferase assays, chromatin immunoprecipitation, electrophoretic mobility shift assays, and quantitative RT-PCR using EGR-1-inducible lung cancer cells demonstrated that EGR-1 binds to the proximal region of the VEGF-A promoter, activates VEGF-A expression, and enhances hypoxia inducible factor 1alpha (HIF-1alpha)-mediated VEGF-A expression. The EGR-1 modulator, NAB-2, was rapidly induced by increased levels of EGR-1. Pathology samples of human lung adenocarcinomas revealed correlations between EGR-1/HIF-1alpha and VEGF-A expressions and relative elevation of EGR-1 and VEGF-A expression in mutant K-RAS- or EGFR-carrying adenocarcinomas. Both EGR-1 and VEGF-A expression increased as tumors dedifferentiated, whereas HIF-1alpha expression did not. Although weak correlation was found between EGR-1 and NAB-2 expressions on the whole, NAB-2 expression decreased as tumors dedifferentiated, and inhibition of DNA methyltransferase/histone deacetylase increased NAB-2 expression in lung cancer cells despite no epigenetic alteration in the NAB-2 promoter. These findings suggest that EGR-1 plays important roles on VEGF-A expression in lung cancer cells, and epigenetic silencing of transactivator(s) associated with NAB-2 expression might also contribute to upregulate VEGF-A expression.

Insulin-like growth factor binding protein-4 gene silencing in lung adenocarcinomas.

Gene silencing by promoter hypermethylation plays an important role in molecular pathogenesis. We previously reported that insulin-like growth factor (IGF) binding protein-4 (IGFBP-4), which inhibits IGF-dependent growth, is expressed via early growth response-1 (EGR-1) and is often silenced in cultivated lung cancer cells. The purpose of the present study was to clarify clinicopathological factors associated with IGFBP-4 gene silencing in lung adenocarcinomas. Seventy-six surgically resected adenocarcinomas (20 well-, 35 moderately-, and 21 poorly-differentiated) were subjected to methylation-specific polymerase chain reaction (PCR) analysis for EGR-1-binding sites located in the IGFBP-4 promoter and immunohistochemistry for IGFBP-4, EGR-1, and Ki-67. Thirty-two adenocarcinomas (42%) revealed IGFBP-4 promoter hypermethylation, and the severity inversely correlated with the level of IGFBP-4 expression (P < 0.0001) and tumor differentiation (well versus poor, P = 0.0278; well/moderate versus poor, P = 0.0395). Furthermore, there was a negative correlation between Ki-67 labeling index and IGFBP-4 expression (P = 0.0361). These findings suggest that the expression of IGFBP-4 in adenocarcinoma cells in vivo is downregulated by epigenetic silencing in association with tumor differentiation, resulting in disruption of the mechanism of IGFBP-4-mediated growth inhibition.

Down-regulation of FXYD3 expression in human lung cancers: its mechanism and potential role in carcinogenesis.

FXYD3 is a FXYD-containing Na,K-ATPase ion channel regulator first identified as a protein overexpressed in murine breast tumors initiated by oncogenic ras or neu. However, our preliminary study revealed that FXYD3 expression was down-regulated in oncogenic KRAS-transduced airway epithelial cells. This contradiction led us to investigate the role of FXYD3 in carcinogenesis of the lung. FXYD3 mRNA and protein levels were lower in most of the lung cancer cell lines than in either the noncancerous lung tissue or airway epithelial cells. Protein levels were also lower in a considerable proportion of primary lung cancers than in nontumoral airway epithelia; FXYD3 expression levels decreased in parallel with the dedifferentiation process. Also, a somatic point mutation, g55c (D19H), was found in one cell line. Forced expression of the wild-type FXYD3, but not the mutant, restored the well-demarcated distribution of cortical actin in cancer cells that had lost FXYD3 expression, suggesting FXYD3 plays a role in the maintenance of cytoskeletal integrity. However, no association between FXYD3 expression and its promoter's methylation status was observed. Therefore, inactivation of FXYD3 through a gene mutation or unknown mechanism could be one cause of the atypical shapes of cancer cells and play a potential role in the progression of lung cancer.

Down-regulation of DUSP6 expression in lung cancer: its mechanism and potential role in carcinogenesis.

Our preliminary studies revealed that oncogenic KRAS (KRAS/V12) dramatically suppressed the growth of immortalized airway epithelial cells (NHBE-T, with viral antigen-inactivated p53 and RB proteins). This process appeared to be a novel event, different from the so-called premature senescence that is induced by either p53 or RB, suggesting the existence of a novel tumor suppressor that functions downstream of oncogenic KRAS. After a comprehensive search for genes whose expression levels were modulated by KRAS/V12, we focused on DUSP6, a pivotal negative feedback regulator of the RAS-ERK pathway. A dominant-negative DUSP6 mutant, however, failed to rescue KRAS/V12-induced growth suppression, but conferred a stronger anchorage-independent growth activity to the surviving subpopulation of cells generated from KRAS/V12-transduced NHBE-T. DUSP6 expression levels were found to be weaker in most lung cancer cell lines than in NHBE-T, and DUSP6 restoration suppressed cellular growth. In primary lung cancers, DUSP6 expression levels decreased as both growth activity and histological grade of the tumor increased. Loss of heterozygosity of the DUSP6 locus was found in 17.7% of cases and was associated with reduced expression levels. These results suggest that DUSP6 is a growth suppressor whose inactivation could promote the progression of lung cancer. We have here identified an important factor involved in carcinogenesis through a comprehensive search for downstream targets of oncogenic KRAS.

Neuroendocrine cancer-specific up-regulating mechanism of insulin-like growth factor binding protein-2 in small cell lung cancer.

Small cell lung cancer (SCLC) exhibits insulin-like growth factor-dependent growth. SCLC is the most aggressive among known in vivo lung cancers, whereas in vitro growth of SCLC is paradoxically slow as compared with that of non-SCLC (NSCLC). In this study, we demonstrate that SCLC cells overexpress insulin-like growth factor binding protein (IGFBP)-2 via NeuroD, a neuroendocrine cell-specific transcription factor. Chromatin immunoprecipitation, electrophoretic mobility shift, and IGFBP-2 promoter assays all revealed that NeuroD binds to the E-box in the 5'-untranslated region of IGFBP-2. A NeuroD transgene in both airway epithelial and NSCLC cells up-regulated the transcription of IGFBP-2 and retarded cell growth. Recombinant IGFBP-2 repressed the growth of both airway epithelial and NSCLC cells in a dose-dependent manner. A NeuroD-specific small interfering RNA repressed IGFBP-2 expression in SCLC, and neutralization of IGFBP-2 and an IGFBP-2-specific small interfering RNA increased SCLC cell growth. Pathological samples of SCLC also expressed IGFBP-2 abundantly, as compared with NSCLC, and showed only rare (8%) IGFBP-2 promoter methylation, whereas the IGFBP-2 promoter was methylated in 71% of adenocarcinomas and 29% of squamous cell carcinomas. These findings suggest that 1) SCLC has an IGFBP-2 overexpression mechanism distinct from NSCLC, 2) secreted IGFBP-2 contributes to the slow growth of SCLC in vitro, and 3) the epigenetic alterations in the IGFBP-2 promoter contribute to the striking differences in IGFBP-2 expression between SCLC and NSCLC in vivo.

Small cell lung cancer: significance of RB alterations and TTF-1 expression in its carcinogenesis, phenotype, and biology.

Small cell lung cancer (SCLC) exhibits highly aggressive behavior and has a poor prognosis. While numerous investigations have been carried out, the exact mechanism of its carcinogenesis and aggressiveness is still unclear. SCLC is categorized as a neuroendocrine neoplasia and has a genetic profile characterized by universal alterations of the RB and TP53 genes. Epidemiological studies indicate the majority of SCLCs to be caused by smoking and the TP53 mutational pattern to be consistent with that evoked by smoke carcinogens; however, there is no direct evidence that such carcinogens induce alterations to RB in SCLC. While the importance of these alterations in the carcinogenesis of SCLC is strongly suggested, the exact molecular mechanism has been only little elucidated. SCLC cells almost always express mammalian achaete-scute homolog-1 (MASH1) and thyroid transcription factor-1 (TTF-1). MASH1 plays a critical role in neuroendocrine differentiation. TTF-1 is a characteristic marker of distal airway cells and pulmonary adenocarcinomas, but is also expressed in extrapulmonary neuroendocrine cancers. Thus, TTF-1 may well play a significant role in the development of neuroendocrine cancers. Recent studies indicate that the airway stem cell is committed to the neuroendocrine lineage through MASH1 and Notch signaling and that only RB-deleted neuroendocrine cells selectively proliferate in response to E2F3, eventually undergoing transformation to neuroendocrine cancer cells, probably in concert with TP53 gene aberrations. Thus, alterations of both the RB and TP53 genes are central to the carcinogenesis of SCLC, while many other factors including MASH1 and TTF-1 contribute to the development and biological behavior of SCLC.

PROX1 Promotes Secretory Granule Formation in Medullary Thyroid Cancer Cells.

Mechanisms of endocrine secretory granule (SG) formation in thyroid C cells and medullary thyroid cancer (MTC) cells have not been fully elucidated. Here we directly demonstrated that PROX1, a developmental homeobox gene, is transcriptionally involved in SG formation in MTC, which is derived from C cells. Analyses using gene expression databases on web sites revealed that, among thyroid cancer cells, MTC cells specifically and highly express PROX1 as well as several SG-forming molecule genes. Immunohistochemical analyses showed that in vivo MTC and C cells expressed PROX1, although follicular thyroid cancer and papillary thyroid cancer cells, normal follicular cells did not. Knockdown of PROX1 in an MTC cells reduced SGs detected by electron microscopy, and decreased expression of SG-related genes (chromogranin A, chromogranin B, secretogranin II, secretogranin III, synaptophysin, and carboxypeptidase E). Conversely, the introduction of a PROX1 transgene into a papillary thyroid cancer and anaplastic thyroid cancer cells induced the expression of SG-related genes. Reporter assays using the promoter sequence of chromogranin A showed that PROX1 activates the chromogranin A gene in addition to the known regulatory mechanisms, which are mediated via the cAMP response element binding protein and the repressor element 1-silencing transcription factor. Furthermore, chromatin immunoprecipitation-PCR assays demonstrated that PROX1 binds to the transcriptional regulatory element of the chromogranin A gene. In conclusion, PROX1 is an important regulator of endocrine SG formation in MTC cells.

Enhancement of pleural dissemination and lymph node metastasis of intrathoracic lung cancer cells by vascular endothelial growth factors (VEGFs).

The expression of vascular endothelial growth factors (VEGFs) in tumors including lung cancer is considered to be associated with tumor development via capillary and lymph vessel neogenesis. Dissemination of the tumor cells to the pleura or regional lymph nodes is a critical poor prognostic factor for lung cancer patients. To investigate how VEGFs expressed in the intrathoracic infiltrating lung cancer cells participate in disease progression, we established stably VEGF-A-, VEGF-C-, VEGF-D-, VEGF-A and VEGF-C-, and VEGF-A and VEGF-D-expressing large cell lung cancer clones (TKB5/VEGF-A, TKB5/VEGF-C, TKB5/VEGF-D, TKB5/VEGF-A/C, and TKB5/VEGF-A/D), orthotopically inoculated these into the right thoracic cavity (i.t.) of nude mice, and evaluated the subsequent development of lung lesion, pleural effusion, pleural dissemination, and lymph node metastasis. While there were no significant differences either in culture or in subcutaneous tumor cell growth between the empty vector-transfected group (TKB5/empty) and each transfectant, the i.t. model demonstrated significantly different biological properties between the transfectants. TKB5/empty-inoculated mice frequently developed a large tumor on the pleura without pleural effusion, dissemination, or lymph node (LN) metastasis. In contrast, VEGF-A promoted a bloody pleural effusion (6/14), and VEGF-A and VEGF-D frequently generated pleural dissemination (11/14 and 9/11, respectively). Although both VEGF-C and VEGF-D generated LN metastasis (6/10 and 8/11, respectively), the locations of the metastasized LNs were quite different. TKB5/VEGF-C metastasized on the same side of axillary LNs as i.t. (right axillary LNs), whereas TKB5/VEGF-D metastasized to the mediastinal and left axillary and/or cervical LNs. Since the TKB5/VEGF-A/C or TKB5/VEGF-A/D co-transfectants revealed overlapping tumor progression patterns of VEGF-A and VEGF-C or VEGF-D, the metastatic LNs had abundant new capillaries and were larger than those of TKB5/VEGF-C or TKB5/VEGF-D-inoculated mice. Our results clearly demonstrate that VEGF-A secreted from intrathoracic lung cancer cells plays important roles in producing pleural effusion, dissemination, and capillary neogenesis, that VEGF-C is involved in LN metastasis, and VEGF-D in pleural dissemination and LN metastasis. It is most likely, however, that the mechanisms by which VEGF-C promotes LN metastasis are different from those of VEGF-D. The regulation of the expression of VEGFs in intrathoracic lung cancer cells might be a useful therapeutic approach to inhibiting tumor development and improving patient prognosis.

Cancer stem cell: implications in cancer biology and therapy with special reference to lung cancer.

The cancer stem cell (CSC) theory is currently central to the field of cancer research, because it is not only a matter of academic interest but also crucial in cancer therapy. CSCs share a variety of biological properties with normal somatic stem cells in terms of self-renewal, the propagation of differentiated progeny, the expression of specific cell markers and stem cell genes, and the utilization of common signaling pathways and the stem cell niche. However, CSCs differ from normal stem cells in their tumorigenic activity. Thus, CSCs are also termed cancer initiating cells. In this paper, we briefly review hitherto described study results and refer to some excellent review articles to understand the basic properties of CSCs. In addition, we focus upon CSCs of lung cancers, since lung cancer is still increasing in incidence worldwide and remains the leading cause of cancer deaths. Understanding the properties of, and exploring cell markers and signaling pathways specific to, CSCs of lung cancers, will lead to progress in therapy, intervention, and improvement of the prognosis of patients with lung cancer. In the near future, the evaluation of CSCs may be a routine part of practical diagnostic pathology.