Structure-activity relationship studies of tetrahydroquinolone derivatives as GPR41 modulators.

GPR41, a G protein-coupled receptor, serves as a sensor for short-chain fatty acids and plays a crucial role in regulating multiple physiological processes such as the maintenance of metabolic and immune homeostasis. Therefore, the modulation of GPR41 has garnered attention as a potential strategy for the treatment of various disorders. We conducted a structure-activity relationship study on a lead tetrahydroquinolone derivative bearing a 2-(trifluoromethoxy)benzene group that displayed antagonistic activity toward GPR41. Modification of the aryl group attached to the furan moiety revealed that derivatives containing di- or trifluorobenzene, instead of 2-(trifluoromethoxy)benzene, exhibited agonistic activity toward GPR41, comparable with the reported agonistic modulator AR420626. These results suggest that the aryl group plays a pivotal role in regulating the activity of compounds toward GPR41, providing valuable insights for the design of GPR41 modulators.

Association between AUTS2 haplotypes and alcohol dependence in a Japanese population.

OBJECTIVE: Recent genome-wide analysis has indicated that the autism susceptibility candidate 2 (AUTS2) gene is involved in the regulation of alcohol consumption. We hypothesised that AUTS2 might be associated with the development of alcohol dependence. Therefore, in this exploratory study, we compared the genotype and allele frequencies of the polymorphisms rs6943555 and rs9886351 in the AUTS2 gene between patients with alcohol dependence and healthy control subjects living in a Japanese provincial prefecture. We also examined whether or not the haplotypes consisting of these polymorphisms are related to alcohol dependence. METHODS: The subjects of this study consisted of 64 patients with alcohol dependence and 75 unrelated healthy people. The AUTS2 genotypes were determined by the polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) method. RESULTS: No significant differences in the genotype and allele frequencies of the polymorphisms AUTS2 rs6943555 and rs9886351 were found between alcohol dependence and control subjects. On the other hand, the frequencies of the AUTS2 haplotypes were significantly different between them, and the rs6943555 and rs9886351 A-A haplotype was associated with alcohol dependence (p=0.0187). CONCLUSION: This suggests that the rs6943555 and rs9886351 A-A haplotype might affect the vulnerability to alcohol dependence pathogenesis. Further studies are needed to confirm the reproducibility of the results of this study with increased numbers of subjects.

[Association between norepinephrine transporter gene polymorphism and alcohol dependence in Japanese].

Several studies have suggested that the norepinephrine transporter (NET) may play an important role in the pathogenesis of alcohol dependence. Therefore, in this study, we investigated whether the NET gene polymorphism is a susceptibility factor for alcohol dependence in 64 alcoholics and 73 healthy controls. In addition, we examined whether the combination of the NET and serotonin transporter genotypes are associated with alcohol dependence. The NET (1287G/A, -182T/C, and -3081A/T) and serotonin transporter (5-HTT3'UTR) genotypes were determined by the polymerase chain reaction (PCR)--restriction fragment length polymorphism (RFLP) method. No significant differences in genotype and allele frequencies of the NET and serotonin transporter gene polymorphisms were found between alcoholics and controls. The haplotype frequencies of the NET gene polymorphisms were not also significantly different between them. Furthermore, the combination of the NET and serotonin transporter genotypes had not significant effects on alcohol dependence. The present study suggests that the polymorphisms of 1287G/A, -182T/C and -3081A/T in NET gene are not.risk factors in alcohol dependence.

[The relationship between BDNF gene polymorphisms and alcoholics in Japan].

As a help of the mechanism elucidation of alcoholism, we studied the relationship between brain-derived neurotrophic factor (BDNF) rs6265, 270 C/T (ID number has not yet been determined), and rs10835210 gene polymorphisms, which are reported to be related to bipolar disorder, and alcoholics. We genotyped the three polymorphisms in the BDNF gene using polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) in 65 alcoholics and 71 healthy controls. In this study, there was no significant difference in the frequency of rs6265 and 270 C/T polymorphisms between alcoholics and controls (P > 0.05). However, there was a significant difference in the genotype frequency of rs10835210 polymorphism between alcoholics and controls (P < 0.05), in which the CA heterozygote genotype and A allele frequency was higher in alcoholics than in the controls. It suggests the possibility that the BDNF rs10835210 gene polymorphism affects the etiology of alcoholism.

[Motivational interviewing and cognitive behavior therapy for smoking cessation].

The combination therapy of counseling and medication is very important in smoking cessation treatment. Motivational interviewing (MI) and cognitive behavior therapy (CBT) are interventions recommended in two well-recognized guidelines for smoking cessation in the United States. MI is effective for those who do not want to quit smoking; CBT for those who cannot but want to quit smoking, those who repeatedly quit and re-smoke, those who are depressed, and women who are concerned about their weight. However, neither MI nor CBT for smoking cessation is widely practiced in Japan. We describe the nature of, evidence for, and practice of MI and CBT in smoking cessation treatment, hopeful that clinicians and other healthcare staff learn and apply these techniques.

[Relationship between GSK-3 beta -50T/C and DBI +529A/T polymorphisms in Japanese alcoholics].

As a help of the relationship between bipolar disorder and alcoholism, we studied the relationship between GSK-3 beta -50T/C polymorphism, which is reported to the relationship for bipolar disorder, and Japanese alcoholics. And we investigated the relationship between GSK-3 beta -50T/C polymorphism and DBI +529A/T polymorphisms, which is reported to one of the risk factor for alcoholism. We analyzed the GSK-3 beta genotype using a polymerase chain reaction (PCR), and DBI genotype using PCR with confronting two-pair novel primers (PCR-CTPP) in 75 health controls and 64 alcoholics. In this study, there was no significant difference in the frequency of GSK-3 beta -50T/C polymorphism between alcoholics and controls (p = 0.883), and there was no significant difference in the frequency of DBI +529A/T polymorphism (p = 0.131). Also, there was no relationship between GSK-3 beta -50T/C polymorphism and DBI +529A/T allele in alcoholism (p = 0.907). We suggested that bipolar disorder may not be one of the pathogenesis of alcoholism, and that there was no relationship between GSK-3 beta -50T/C polymorphism and DBI +529A/T polymorphism.

[The smoking situation of patients and staff in a psychiatry hospital. The smoking issue in psychiatry can be considered a neglected problem].

OBJECTIVE: The present study was conducted to obtain information about smoking status of psychiatric patients and to determine whether there might be a demand for smoking cessation support for this group of people. In addition, the smoking status of psychiatric medical staff members, their awareness regarding smoking and related issues, and their attitude to promotion efforts to ameliorate smoking in their working place were examined. METHODS: Outpatients and inpatients with schizophrenia, mood disorders, and alcoholism in Koutokukai Sato Hospital during December 2001 and May 2002, and staff of the hospital were the subjects in this study. We surveyed smoking status in both 296 patients and 222 staff members. RESULTS: Smoking rates were 77.4% in males and 39.3% in females among patients with schizophrenia, 87.5% in males and 100% in females among patients with bipolar mood disorders, 69.6% in males and 5.4% in females among patients with depression, and 86.7% in males and 100% in females among patients with alcoholism. Among those smokers, 78.1% were nicotine dependent. However, 75.7% of these smokers were interested in smoking cessation, and 49.0% hoped for prohibition of smoking. The findings thus indicated that the demand for smoking cessation support is high in psychiatric patients. Among the staff, the smoking rate was also high, at 45.5% (males: 76.6% and females: 29.0%). As for the age at beginning of smoking, the peaks were at 18 years old and 20 years old. Smokers who smoked less than 20 cigarettes per day accounted for 80% of the total. Of the smokers, 91.1% recognized that their smoking bothered the people around them. If the working place was smoke free, however, 66.3% answered it would be difficult to adapt, and only 24% wanted to stop smoking recently. On the other hand, 29.8% of the non-smokers were bothered with smoking at the working place, and 76.0% hated smoke of cigarettes. When one wanted a smoker not to smoke, 22.7% of the non-smokers could say so. Of the staff members, 80.0% agreed with anti-smoking measures. However, it appeared that their consciousness of smoking issues was low as medical workers. CONCLUSIONS: The smoking rate of psychiatric patients and the prevalence of nicotine dependence are high. However, half of the subjects in the present study expressed a desire to quit smoking, pointing to a high demand of smoking cessation support. Psychiatric staff at the institution studied had a high smoking rate, and their recognition of smoking issues was low.

Need for medication to complement catecholamines in smoking cessation of hardcore smokers.

Many smokers find it difficult to stop smoking without assistance. The antidepressants bupropion and nortriptyline can aid smoking cessation. The main aim of this study was to understand the pathophysiology of smoking cessation better based on biological backgrounds. We investigated the following biological markers for any alterations during smoking cessation in the absence of pharmacotherapy: the dopamine metabolite homovanillic acid (HVA), the noradrenaline metabolite 3-methoxy-4-hydroxyphenylglycol (MHPG) and brain-derived neurotrophic factor (BDNF). Assessment and blood sampling were performed at a baseline (the start) time point and at a critical time point during smoking cessation. Seven of 30 smokers quit during a 16-week follow-up period; these smokers were defined as remission group from tobacco dependence. The remaining 23 smokers were categorized as hardcore smokers. The smoking group was compared with 23 non-smokers matched for age and gender. We compared blood levels of biological markers in each of the three groups. The hardcore smoker group showed significant decreases in HVA and MHPG levels between baseline and the critical time point (p=0.018 and p=0.033, respectively). However, the remission from tobacco dependence group exhibited no significant changes in any of the biomarkers examined. They had lower scores on the Minnesota nicotine withdrawal scale than the hardcore smoker group (p=0.002). The hardcore smoker group had higher MHPG and BDNF levels than the non-smoker group (p=0.002 and p<0.001, respectively). Hardcore smokers experience severe nicotine withdrawal symptoms. Nicotine withdrawal is associated with catecholamine deficiency. The resulting withdrawal symptoms make quitting difficult for hardcore smokers. These hardcore smokers may require medication to compensate for the catecholamine deficit. Non-nicotinic medications such as bupropion, nortriptyline, or varenicline may be required to bolster the catecholamine deficit in hardcore smokers.

Continuous in vivo culture and indirect fluorescent antibody test for zoonotic protozoa of Babesia microti.

Babesiosis has attracted attention as a zoonotic disease. The disease is caused in immunocompetent individuals almost solely by Babesia microti, a rodent babesia. Most cases of human babesiosis by B. microti have been reported in the endemic foci of the Northeastern coastal areas and upper Midwest regions of the United States, while some sporadic cases have recently been reported in several Asian countries including Japan. Our previous surveys identified that four small subunit ribosomal RNA gene (SSUrDNA) types of B. microti parasitize Japanese rodents. Indirect fluorescent antibody test (IFAT) is often performed for the diagnosis of babesiosis together with microscopical examination of thin blood smears and PCR. We established IFAT against four SSUrDNA-types of B. microti using erythrocytes of SCID mice or Syrian hamsters infected with each SSUrDNA-type B. microti. The results of IFAT for sera of ICR mice or Syrian hamsters infected with each SSUrDNA-type B. microti demonstrated that the four SSUrDNA-types of B. microti have different serotypes. Here, we report technical or practical procedures of IFAT, which gains sufficiently stable results, including procedures of continuous in vivo culture of B. microti.

Development of real-time PCR assay for differential detection and quantification for multiple Babesia microti-genotypes.

We have developed a real-time PCR assay that can rapidly and differentially detect and quantify four genotypes of small subunit ribosomal RNA gene (SSUrDNA) of Babesia microti (Kobe-, Otsu-, Nagano- and US-types). In this assay, four genotype-specific pairs of primers targeted on internal transcribed spacer (ITS) 1 or 2 sequences were used and amplicons by each pair of primers were quantitatively detected by fluorescent SYBR Green I. The four genotype-specific pairs of primers displayed the high specificity for homologous genotype DNA. The standard curves of cycle threshold (Ct) values versus amount of target DNA per reaction (log) for all four genotypes were linear and the correlation coefficient (Rsq) values for the curves were from 0.970 to 0.997. The standard curves were almost identical even in the presence of heterologous genotype DNA. This assay could detect 10-30 fg purified DNA (equivalent to the amount of 1-5 parasite DNA) of each genotype B. microti. This assay could also detect each genotype B. microti infection in blood with 3×10(-6)%-1×10(-5)% parasitemia. This assay was applicable to field rodent and tick samples to reveal mixed infection in several samples, for which a single genotype of B. microti had been detected by direct sequencing analyses in our previous studies. This assay also seemed to be applicable to clinical human samples, showing Kobe-type positive results for the first Japanese babesiosis patient and the asymptomatic donor, both infected with Kobe-type B. microti.

Efficacy of chloroquine plus primaquine treatment and pfcrt mutation in uncomplicated falciparum malaria patients in Rangamati, Bangladesh.

A combination of chloroquine (CQ) and primaquine (PQ) had been used as the first-line treatment of uncomplicated Plasmodium falciparum malaria in Rangamati, Bangladesh until the end of 2004. Doctors or medical staffs had felt that CQ plus PQ had become less effective against uncomplicated falciparum malaria patients, but that it was more effective against the minority-indigenous patients than the Bengali patients. The efficacy of CQ plus PQ and the mutation status of the CQ resistance transporter (pfcrt) gene of infecting P. falciparum were, thus, investigated for 45 uncomplicated falciparum malaria patients in Rangamati in 2004. The total failure rate was 57.8%. One or two pfcrt sequences (CIETH and SMNTH at positions 72, 74-76, and 97, mutation underlined) with K76T mutation known to be related to CQ-resistant phenotype were detected in 38 patients' blood samples. Of the 38 patients, in total 15 patients (14/25 minority-indigenous and 1/13 Bengali patients) resulted in adequate clinical and parasitological response (ACPR). There was a statistically significant difference in ACPR rate between the minority-indigenous patients and the Bengali patients. P. falciparum with mutant or resistant pfcrt (pfcrt-resistant) was detected by PCR in blood samples on day 28 for 10 ACPR minority-indigenous patients but not for the only one Bengali ACPR patient, who all were infected with pfcrt-resistant P. falciparum on day 0. The minority-indigenous patients, but not Bengalis, are suggested to be often cured by CQ plus PQ, leaving a very few parasites detectable only by PCR, even when they are infected with pfcrt-resistant P. falciparum.

Survey of Babesia microti infection in field rodents in Japan: records of the Kobe-type in new foci and findings of a new type related to the Otsu-type.

Of 247 rodents comprising 5 genera and 7 species collected at 17 sites throughout Japan from 2003 to 2005, Babesia microti was detected microscopically and by polymerase chain reaction (PCR) in 36 rodents comprising 2 genera and 3 species from 12 sites. Based on the analysis of small subunit ribosomal RNA gene (SSUrDNA) sequences, the Kobe-type, the etiological type of the first Japanese case of human infection was found in Apodemus speciosus and Apodemus argenteus in Aomori, the northernmost prefecture of the Japanese mainland, while the U.S.-type was found on Hokkaido Island and the Otsu-type was widely distributed. In addition, a new Otsu-related type was detected exclusively in Eothenomys andersoni in Nagano, a prefecture in central Japan. The sequences of internal transcribed spacer 1 to 2 (ITS1/2) of the present Kobe- and Otsu-types were almost identical to those of the same types previously identified. The ITS1/2 sequence of the U.S.-type identified in Hokkaido in this survey was somewhat different from that of the U.S.-type strain originating from the U.S.A., with approximately 95% identity. This value was similar to the 94% identity found between the ITS1/2 sequences of the Otsu-type and the new Otsu-related type. The new Otsu-related type of B. microti was isolated as the Nagano strain, which was serologically differentiated from the other type strains of B. microti. The divergence and distribution of genotypes are important factors in investigating the epidemiology of human B. microti infection in Japan.

Association of mu-opioid receptor gene polymorphism A118G with alcohol dependence in a Japanese population.

Ethanol is considered to activate the brain reward system by increasing the release of an endogenous opioid receptor ligand, beta-endorphin. The polymorphism A118G in the mu-opioid receptor gene (OPRM1) causes the amino acid change Asn40Asp and has been reported to affect the affinity of the ligand for the receptor. The association of this polymorphism with the vulnerability to alcohol dependence has been studied in many populations, but not yet in Japanese people. In the present study, we compared the frequencies of the polymorphism OPRM1 A118G between patients with alcohol dependence and healthy control subjects living in a Japanese provincial prefecture. We also genotyped a polymorphism, G1510A, in the acetaldehyde dehydrogenase 2 gene (ALDH2), in which the A allele causes poor metabolism of acetaldehyde, a major metabolite of alcohol. Both OPRM1 118G and ALDH2 1510G were significantly associated with alcohol dependence. These results suggest that OPRM1 118G in addition to ALDH2 1510G might be one of the risk factors for alcohol dependence in Japanese people.