Midkine Promotes Odontoblast-like Differentiation and Tertiary Dentin Formation.

Autophagy is an intracellular self-degradation process that is essential for tissue development, cell differentiation, and survival. Nevertheless, the role of autophagy in tooth development has not been definitively identified. The goal of this study was to investigate how autophagy is involved in midkine (MK)-mediated odontoblast-like differentiation, mineralization, and tertiary dentin formation in a mouse tooth pulp exposure model. In vitro studies show that MK and LC3 have similar expression patterns during odontoblast-like cell differentiation. Odontoblast-like cell differentiation is promoted through MK-mediated autophagy, which leads to increased mineralized nodule formation. Subcutaneous transplantation of hydroxyapatite/tricalcium phosphate with rMK-treated human dental pulp cells led to dentin pulp-like tissue formation through MK-mediated autophagy. Furthermore, MK-mediated autophagy induces differentiation of dental pulp cells into odontoblast-like cells that form DSP-positive tertiary dentin in vivo. Our findings may provide 1) novel insight into the role of MK in regulating odontoblast-like differentiation and dentin formation in particular via autophagy and 2) potential application of MK in vital pulp therapy.

PIN1 is a new therapeutic target of craniosynostosis.

Gain-of-function mutations in fibroblast growth factor receptors (FGFRs) cause congenital skeletal anomalies, including craniosynostosis (CS), which is characterized by the premature closure of craniofacial sutures. Apert syndrome (AS) is one of the severest forms of CS, and the only treatment is surgical expansion of prematurely fused sutures in infants. Previously, we demonstrated that the prolyl isomerase peptidyl-prolyl cis-trans isomerase interacting 1 (PIN1) plays a critical role in mediating FGFR signaling and that Pin1+/- mice exhibit delayed closure of cranial sutures. In this study, using both genetic and pharmacological approaches, we tested whether PIN1 modulation could be used as a therapeutic regimen against AS. In the genetic approach, we crossbred Fgfr2S252W/+, a mouse model of AS, and Pin1+/- mice. Downregulation of Pin1 gene dosage attenuated premature cranial suture closure and other phenotypes of AS in Fgfr2S252W/+ mutant mice. In the pharmacological approach, we intraperitoneally administered juglone, a PIN1 enzyme inhibitor, to pregnant Fgfr2S252W/+ mutant mice and found that this treatment successfully interrupted fetal development of AS phenotypes. Primary cultured osteoblasts from Fgfr2S252W/+ mutant mice expressed high levels of FGFR2 downstream target genes, but this phenotype was attenuated by PIN1 inhibition. Post-translational stabilization and activation of Runt-related transcription factor 2 (RUNX2) in Fgfr2S252W/+ osteoblasts were also attenuated by PIN1 inhibition. Based on these observations, we conclude that PIN1 enzyme activity is important for FGFR2-induced RUNX2 activation and craniofacial suture morphogenesis. Moreover, these findings highlight that juglone or other PIN1 inhibitors represent viable alternatives to surgical intervention for treatment of CS and other hyperostotic diseases.

Arsenic exposure and seafood intake in Korean adults.

Arsenic (As) is widely distributed in the environment, and humans can be exposed to As from various sources such as air, water, soil, and food. This study was performed to evaluate the As exposure levels in Korean adults by measuring total As in urine and its relation with the consumption of seafood, a favorite food in Korea. A total of 2077 adults were the study subjects; they ranged in age from 19 to 83, and they were recruited by probability sampling stratified by area, sex, and age. None of the subjects had been exposed to As occupationally. We collected information about the demographic characteristics, lifestyles, and food consumption of study subjects using a questionnaire and followed urine sampling. Diet was assessed in individual interviews using the 24-h recall method. Total As in urine was analyzed using inductively coupled plasma mass spectrometry (PerkinElmer NEXION 300S; Concord, Ontario, Canada). The geometric mean concentration of total As in urine was observed to be 97.6 µg/L and was higher in males (103.9 µg/L) than in females (93.0 µg/L). Total As levels in urine were affected by sex, age, seafood intake, and geographic location. In this study, total As in urine was positively correlated with fish and shellfish consumption, and was mainly determined by As intake through fish and shellfish/grains/flavors. These findings suggest that seafood consumption might be a major contributor to urinary As levels in Korean adults.

Paratubal cancer found at the time of laparoscopic surgery for adnexal torsion: a case report and literature review.

INTRODUCTION: Paratubal cysts are common incidental finding, but malignant paratubal cancers have rare occurrence and have not been sufficiently described and discussed in previous studies. CASE REPORT: This report describes a case of a 70-year-old female who underwent emergent laparoscopy for adnexal torsion. A serous cystadenocarcinoma arising in a paratubal cyst and accompanied by tubal torsion was revealed by frozen section and successfully treated with laparoscopic cytoreductive surgery and adjuvant chemotherapy. CONCLUSION: This report is the first case of paratubal cancer with bilateral tubal torsion which was diagnosed and treated with laparoscopic surgery, and the third report describing serous cystadenocarcinoma arising in a paratubal cyst. In the laparoscopic surgery for the paratubal cyst clinically presumed as accompanied with tubal torsion, surgeons should not ignore the possibility of malignancy in spite of the rare incidence of paratubal cancers and the preconception that adnexal malignancies are seldom accompanied by tubal torsion.

Nestin expression as an indicator of cervical cancer initiation.

Nestin is an intermediate filament protein expressed in proliferating cells during embryonic development of the central nervous system (CNS) and considered to be a neuronal stem cell/progenitor cell marker. This study investigated the difference of nestin expression between pre-cancer (carcinoma in situ - CIS) and cancer of cervix in 129 tissues (49 normal cervix, 41 CIS, and 39 invasive cervical cancer) through the use of a paraffin-embedded tissue array. Immunostaining was evaluated by intensity, proportion of stained cells, and pattern of expression. The expression of nestin was positive in 63.4% (26/41) for CIS and 43.6% (17/39) for invasive cervical cancer, but only 26.5% (13/49) for normal tissues (p = 0.002). Strong positive staining/large proportion staining were 53.7% (22/41) / 36.6% (15/41), 15.4% (6/39) / 61.5% (24/39) in the CIS and invasive cervical cancer tissues, respectively (p = 0.043, p < 0.001). The diffuse stain with basal layer was positive in 90.2% (37/41) for CIS, but only 24.5% (12/49) of the samples were positive in normal tissues (p < 0.001). Based on these results, the authors suggest that nestin expression seems to participate in the step of cancer initiation and could potentially be a useful marker in the early detection of cervical cancer.

Demographic and serological characteristics of Asian Americans with hepatitis B infection diagnosed at community screenings.

There is limited information regarding follow-up and hepatitis B serological status of Asian Americans diagnosed with chronic hepatitis B (CHB) through community screening. The aims of this study were to evaluate the prevalence and characterize CHB among Asians living in Los Angeles, assess follow-up of individuals with CHB diagnosed at screening and compare with patients with CHB followed by community gastroenterologists. Between October 2007 and May 2010, 7387 Asians were tested for HBV. HBsAg positive individuals (CHB) underwent additional testing for ALT, HBeAg/anti-HBe and HBV DNA. Patients with CHB were contacted 6 months later to determine whether they received follow-up care. We compared serological patterns of these individuals with CHB to patients with CHB who were seen for the first time (treatment naïve) by community gastroenterologists during the study period. Prevalence of CHB was 5.2%. About 99% patients with CHB were foreign-born, and only 27% could read/write English. 297 (77%) patients with CHB could be reached 6 months after diagnosis; 43% did not receive follow-up care, mostly because of lack of medical insurance. Patients with CHB followed by gastroenterologists were more likely to have insurance (69% vs 26%, P < 0.0001). 90% patients with CHB at screening were HBeAg negative/anti-HBe positive with 62% having inactive disease compared to only 30% of patients seen by gastroenterologists (P < 0.0001). Among CHB participants, 13% met criteria for treatment compared to 51% of patients with CHB (P < 0.0001). Only a small number of CHB screening participants require antiviral therapy. Lack of medical insurance is the main reason for most patients with CHB not seeking follow-up care after screening.

Clostridium chromiireducens sp. nov., isolated from Cr(VI)-contaminated soil.

A Cr(VI)-resistant, Gram-positive, spore-forming, obligate anaerobe, designated GCAF-1(T), was isolated from chromium-contaminated soil by its ability to reduce Cr(VI) in low concentrations. Mixed acid fermentation during growth on glucose resulted in accumulation of acetate, butyrate, formate and lactate. Morphological studies indicated the presence of peritrichous flagella, pili and an S-layer. The major cellular fatty acids (>5 %) were C(16 : 0), C(14 : 0), summed feature 3 (comprising iso-C(15 : 0) 2-OH and/or C(16 : 1)ω7c), C(18 : 1)ω7c, C(16 : 1)ω9c, summed feature 4 (comprising iso-C(17 : 1) I and/or anteiso-C(17 : 1) B) and C(18 : 1)ω9c. The DNA G+C content of strain GCAF-1(T) was 30.7 mol%. Phylogenetic interference indicated that strain GCAF-1(T) clustered with group I of the genus Clostridium. Of strains within this cluster, strain GCAF-1(T) shared the highest 16S rRNA gene sequence similarities (98.1-98.9 %) with Clostridium beijerinckii DSM 791(T), C. saccharobutylicum NCP 262(T), C. saccharoperbutylacetonicum N1-4(T), C. puniceum DSM 2619(T) and C. roseum DSM 51(T). However, strain GCAF-1(T) could be clearly distinguished from its closest phylogenetic neighbours by low levels of DNA-DNA relatedness (<50 %) and some phenotypic features. Based on the evidence presented here, strain GCAF-1(T) ( = DSM 23318(T) = KCTC 5935(T)) represents a novel species of the genus Clostridium, for which the name Clostridium chromiireducens sp. nov. is proposed.

Modulation of natural killer cell activity affected by electroacupuncture through lateral hypothalamic area in rats.

Electroacupuncture (EA) has been reported to modulate natural killer cell (NK cell) activities. Also it is well known that hypothalamus directly mediates the effects of EA on analgesia. Especially lateral hypothalamic area (LHA) is related to splenic NK cell activities. In order to investigate the relationship between hypothalamus and effects of EA on NK cell activity, lesions have been made bilaterally at LHA of Spraque-Dawley rats. Subsequently, NK cell cytotoxities of normal and lesioned rats were measured with (51)Cr release immunoassay after EA stimulation for 2 and 14 days. NK cell activity of EA group was significantly higher than sham group. In addition, lesions abolished effects of EA on NK cell activity. These results strongly suggest that LHA is closely related to increase of NK cell activity induced by EA.

Developing and sustaining 3-chlorophenol-degrading populations in up-flow anaerobic column reactors under circum-denitrifying conditions.

Microbial consortia capable of degrading 3-chlorophenol (3-CP) were enriched in continuous up-flow column reactors under circum-denitrifying conditions. 3-CP degradation capability was developed and sustained when 3-CP was supplied at 16-21 microM, although suppression of the 3-CP degradation capability was observed when 3-CP was supplied at 42 microM. When 3-CP was stably degraded, the ratio of nitrate consumption to 3-CP degradation approached the theoretical stoichiometric value, which was calculated by assuming a 3-CP degradation-dependent nitrate reduction. Batch-incubation experiments demonstrated that the microbial consortium that was enriched in the column reactors required either nitrate or oxygen for degrading 3-CP, while 3-CP was not degraded under sulfate-degrading conditions. Although many attempts were made to sustain the microbial 3-CP degradation capability under denitrifying conditions, mostly in batch cultures, none of them have been successful so far. Therefore, the results obtained in this study may be the first to demonstrate sustainable 3-CP degradation capability under circum-denitrifying conditions.

A controlled clinical study of serosa-invasive gastric carcinoma patients who underwent surgery plus intraperitoneal hyperthermo-chemo-perfusion (IHCP).

BACKGROUND: Despite recent advances in the treatment of advanced gastric carcinomas, no satisfactory outcomes are available because of micrometastases and free-floating carcinoma cells already existing in the peritoneal cavity. From 1990, we started using intraperitoneal hyperthermo-chemo-perfusion (IHCP) to prevent and to treat peritoneal metastasis after surgical resection of stomach cancer. METHOD: We analyzed 103 serosa-invasive gastric carcinoma patients who underwent surgical resection between 1990 and 1995. Fifty-two patients who received surgery plus IHCP were compared with 51 patients who underwent surgery only, as controls. IHCP was administered for 2 h with an automatic IHCP device (closed-circuit system) just after surgical resection, with the patient under hypothermic general anesthesia (32.4 degrees C-34.0 degrees C). As perfusate, we used 1.5% peritoneal dialysis solution mixed with 10 micrograms/ml of mitomycin-C (MMC), warmed at an inflow temperature of over 44 degrees C. RESULTS: The overall 5-year survival rate (5-YSR) of the 103 patients was 29.97%. The 5-YSR was higher in the IHCP group than in the control group, at 32.7% and 27.1%, respectively, but this difference was not significant. However, in the 65 serosa-invasive gastric carcinoma patients (excluding those in stage IV) the 5-YSR was significantly higher (P = 0.0379) in the IHCP group than in the control group, at 58.6% and 44.4%, respectively. On multivariate analysis of all 103 patients, depth of tumor invasion and lymph node metastasis were significant factors for survival, whereas significant factors on univariate analysis, such as combined operation, distant metastasis, and peritoneal metastasis, were not significant. The most common recurrence patterns were loco-regional in the IHCP group and peritoneal in the control group. CONCLUSION: Complete cytoreductive surgery plus IHCP is effective to prevent and to treat peritoneal metastasis, and it should lead to long-term survival for serosa-invasive gastric carcinoma patients. However, there are some limitations for the treatment of peritoneal metastasis in stage IV patients with distant metastases.

A novel denitrifying bacterial isolate that degrades trimethylamine both aerobically and anaerobically via two different pathways.

The aerobic and anaerobic degradation of trimethylamine by a newly isolated denitrifying bacterium from an enrichment culture with trimethylamine inoculated with activated sludge was studied. Based on 16S rDNA analysis, this strain was identified as a Paracoccus sp. The isolate, strain T231, aerobically degraded trimethylamine, dimethylamine and methylamine and released a stoichiometric amount of ammonium ion into the culture fluid as a metabolic product, indicating that these methylated amines were completely degraded to formaldehyde and ammonia. The strain degraded trimethylamine also under denitrifying conditions and consumed a stoichiometric amount of nitrate, demonstrating that complete degradation of trimethylamine was coupled with nitrate reduction. Cell-free extract prepared from cells grown aerobically on trimethylamine exhibited activities of trimethylamine mono-oxygenase, trimethylamine N-oxide demethylase, dimethylamine mono-oxygenase, and methylamine mono-oxygenase. Cell-free extract from cells grown anaerobically on trimethylamine and nitrate exhibited activities of trimethylamine dehydrogenase and dimethylamine dehydrogenase. These results indicate that strain T231 had two different pathways for aerobic and anaerobic degradation of trimethylamine. This is a new feature for trimethylamine metabolism in denitrifying bacteria.

Association between TNF-alpha promoter polymorphism and Helicobacter pylori cagA subtype infection.

AIMS: To assess the importance of tumour necrosis factor alpha (TNF-alpha) promoter polymorphism in relation to infection with the cytotoxin associated gene A (cagA) subtype of Helicobacter pylori within a dyspeptic Korean population. METHODS: Eighty three patients with gastric disease and 113 healthy controls were studied. The DNA from gastric biopsy specimens was analysed by H pylori specific and cagA specific polymerase chain reaction (PCR). To characterise TNF-alpha polymorphism at positions -308 and -238, PCR based restriction fragment length polymorphism analysis was performed. RESULTS: Helicobacter pylori infection was closely correlated with G to A transition at position -308 of the TNF-alpha promoter when compared with healthy controls (odds ratio (OR), 2.912; 95% confidence interval (CI), 1.082 to 7.836; p = 0.034). Although TNF-alpha -308 polymorphism in patients with H pylori was not significantly different from that in patients without H pylori, the -308A polymorphism was strongly associated with H pylori cagA subtype infection when compared with the polymorphism in cagA negative H pylori infection (OR, 8.757; 95% CI, 1.413 to 54.262; p = 0.019) and healthy controls (OR, 3.683; 95% CI, 1.343 to 10.101; p = 0.011). G to A genetic change at position -238 of the TNF-alpha gene was not significantly associated with H pylori cagA subtype infection. In addition, genetic polymorphisms at both sites of the TNF-alpha promoter in patients with H pylori infection did not correlate with the severity of disease. CONCLUSION: TNF-alpha -308A polymorphism was significantly related to infection with the H pylori cagA subtype in Korean patients with gastric disease.

Effects of uwhangchungsimwon on cell viability, proliferation, and gene expression of human neuronal cell line IMR32.

Uwhangchungsimwon (pill, UC) is one of the traditional Korean medical prescriptions that has been most frequently used for stroke. To characterize the effects of UC on human neuronal cells, the human neuroblastoma cell line IMR32 was treated with UC, and cell viability, cell proliferation, apoptosis, and gene expression were analyzed. The effect of UC on recovery of cell viability was analyzed following stress induction by nutrient depletion or cold shock. Flow cytometric analysis of the cell cycle showed that UC inhibits cell cycle progression of IMR32 in a dose- and time-dependent manner. UC was also identified to increase cell viability and suppress apoptosis induction by a DNA-damaging agent, etoposide. Quantitative RT-PCR analysis revealed that expressions of the p53 tumor suppressor gene and its downstream effect, Waf1, are stimulated whereas expressions of positive cell cycle regulators, c-Myc, c-Fos, and Cyclin D1 were repressed by UC treatment. Moreover, while expression levels of apoptosis inhibitors, Bcl-2 and Bcl-XL were increased following UC treatment, that of an apoptosis promoter, Bax, was decreased. In addition, expression of BMP-7, which has been recently demonstrated to improve the motor neuron recovery from stroke, was induced by UC while it was not detected in untreated cells. Taken together, our data suggest that the pharmacoclinical effects of UC might be derived in part from its negative regulation of cell proliferation and apoptosis through the transcriptional control of related genes.

Inhibition of human smooth muscle cell proliferation by gamigeonsim-tang through the transcriptional regulation of cell cycle-controlling genes.

The effects of gamigeonsim-tang (GGT) on cellular proliferation and expression of cell cycle-related genes were investigated in human smooth muscle cell HISM. HISM cells were treated with an aqueous extract of GGT. Cellular proliferation was investigated by an immunocytometric analysis of PCNA expression and a flow cytometric analysis of the cell cycle progression. Reduced expression of PCNA and a significant accumulation of G1 phase cells were observed following treatment, indicating that GGT inhibits cellular proliferation of human smooth muscle cells. To explore whether GGT affects the transcription of cell cycle-regulating genes, we evaluated mRNA expression of p53, p21Waf1 PCNA, Cyclin D1, Cdc2, Histone H3, c-Myc, and c-Fos using a quantitative RT-PCR analysis. While increased expressions of two negative cell cycle regulators, p53 and p21Waf1 were found, reduced expressions of cell cycle stimulators, PCNA, c-Fos, and c-Myc, were identified following treatment. Taken together, our study demonstrates that GGT inhibits cellular proliferation of human smooth muscle cell through the up- and down-regulation of growth-inhibiting and growth-promoting genes, respectively.

A phase II study of VP-16-fosfamide-cisplatin combination chemotherapy plus early concurrent thoracic irradiation for previously untreated limited small cell lung cancer.

BACKGROUND: At present the addition of thoracic irradiation to combination chemotherapy is a standard treatment for limited staged small cell lung cancer. However, there is still controversy about the optimum timing of chest irradiation. We conducted a phase II study of etoposide (VP-16)-ifosfamide-cisplatin (VIP) combination chemotherapy plus early concurrent thoracic irradiation for the patients with previously untreated limited small cell lung cancer in order to assess if the treatment modality could improve the response rate and the toxicity. METHODS: Forty-four patients with limited small cell lung cancer were treated with etoposide-ifosfamide-cisplatin and concurrent thoracic irradiation. Combination chemotherapy consisted of etoposide 100 mg/m2 (on days 1-3), ifosfamide 1000 mg/m2 (on days 1 and 2) and cisplatin 100 mg/m2 (on day 1). Concurrent thoracic irradiation consisted of a total of 4000 cGy over 4 weeks starting on the first day of the first chemotherapy. All patients who showed a complete response were given prophylactic cranial irradiation for 2.5 weeks. RESULTS: Forty-four of the 49 patients who entered the study from May 1994 to August 1998 were evaluable. The median age was 59 years and 40 patients had a performance status of 0 or 1. The median survival time was 22.5 months. Twenty-eight patients (62%) showed a complete response and 16 (38%) a partial response. Twenty-four patients (54%) developed grade 3 or 4 neutropenia; there was a 9% RTOG score 3 or 4 esophagitis. CONCLUSION: VIP combination chemotherapy and early concurrent thoracic irradiation for patients with limited stage small cell lung cancer revealed excellent antitumor response with tolerable toxicity.

Anaerobic and aerobic degradation of pyridine by a newly isolated denitrifying bacterium.

New denitrifying bacteria that could degrade pyridine under both aerobic and anaerobic conditions were isolated from industrial wastewater. The successful enrichment and isolation of these strains required selenite as a trace element. These isolates appeared to be closely related to Azoarcus species according to the results of 16S rRNA sequence analysis. An isolated strain, pF6, metabolized pyridine through the same pathway under both aerobic and anaerobic conditions. Since pyridine induced NAD-linked glutarate-dialdehyde dehydrogenase and isocitratase activities, it is likely that the mechanism of pyridine degradation in strain pF6 involves N-C-2 ring cleavage. Strain pF6 could degrade pyridine in the presence of nitrate, nitrite, and nitrous oxide as electron acceptors. In a batch culture with 6 mM nitrate, degradation of pyridine and denitrification were not sensitively affected by the redox potential, which gradually decreased from 150 to -200 mV. In a batch culture with the nitrate concentration higher than 6 mM, nitrite transiently accumulated during denitrification significantly inhibited cell growth and pyridine degradation. Growth yield on pyridine decreased slightly under denitrifying conditions from that under aerobic conditions. Furthermore, when the pyridine concentration used was above 12 mM, the specific growth rate under denitrifying conditions was higher than that under aerobic conditions. Considering these characteristics, a newly isolated denitrifying bacterium, strain pF6, has advantages over strictly aerobic bacteria in field applications.

Biodegradation of 4-chlorophenol via a hydroquinone pathway by Arthrobacter ureafaciens CPR706.

A newly isolated Arthrobacter ureafaciens, strain CPR706, could degrade 4-chlorophenol via a new pathway, in which the chloro-substituent was eliminated in the first step and hydroquinone was produced as a transient intermediate. Strain CPR706 exhibited much higher substrate tolerance and degradation rate than other strains that degraded 4-chlorophenol by the hydroxylation at the second carbon position to form chlorocatechol. Strain CPR706 could also degrade other para-substituted phenols (4-nitro-, 4-bromo-, 4-iodo-, and 4-fluoro-phenol) via the hydroquinone pathway.