Effects of a laughter program on body weight and mental health among Japanese people with metabolic syndrome risk factors: a randomized controlled trial.

BACKGROUND: While there have been several intervention studies on the psychological effects of laughter, few have examined both the psychological and physical effects. This study investigates the effects of a laughter program on body weight, body mass index (BMI), subjective stress, depression, and health-related quality of life (HRQOL) among Japanese community-dwelling individuals using a randomized controlled trial with a waitlist. METHODS: Overall, 235 participants (37 men and 198 women) aged 43-79 years (mean 66.9, median 67.0) were randomized into laughter intervention and control groups (n = 117 and n = 118, respectively) to participate in a 12-week laughter program. Body weight, subjective stress, subjective well-being, and HRQOL were measured at the baseline, with a 12-week follow-up. The laughter program intervention's effects on these factors were analyzed using an analysis of covariance adjusted by age, sex, risk factors, medication, and area. Furthermore, Pearson's correlation and a general linear model analyzed the relationship between participants' BMI and psychological index changes. RESULTS: The comprehensive laughter program significantly improved the mean body weight (p = 0.008), BMI (p = 0.006), subjective stress (p = 0.004), subjective well-being (p = 0.002), optimism (p = 0.03), and physical component summary (PCS) scores of HRQOL (p = 0.04). A similar tendency occurred for the mean changes in BMI and subjective stress score by area, sex, and age. Moreover, there was a significant and negative correlation between the change in BMI and PCS change (p = 0.04). CONCLUSION: The comprehensive 12-week laughter intervention program, mainly comprising laughter yoga, significantly improved physical and psychological functions such as body weight, BMI, subjective stress, subjective well-being, and HRQOL among predominantly elderly Japanese community-dwelling individuals with metabolic syndrome risk factors. Moreover, PCS improved among participants who reduced BMI after the intervention. These results suggest that the laughter program may help reduce body weight in participants with metabolic syndrome risk factors by reducing stress and improving HRQOL and mental health factors, such as subjective well-being and optimism. TRIAL REGISTRATION: Registered with the University Hospital Medical Information Network Clinical Trials Registry UMIN-CTR000027145 on 27/04/2017.

Working Hours and Risk of Acute Myocardial Infarction and Stroke Among Middle-Aged Japanese Men　- The Japan Public Health Center-Based Prospective Study Cohort II.

BACKGROUND: Evidence from prospective cohort studies regarding the relationship between working hours and risk of cardiovascular disease is limited Methods and Results: The Japan Public Health Center-Based Prospective Study Cohort II involved 15,277 men aged 40-59 years at the baseline survey in 1993. Respondents were followed up until 2012. During the median 20 years of follow up (257,229 person-years), we observed 212 cases of acute myocardial infarction and 745 stroke events. Cox proportional hazards models adjusted for sociodemographic factors, cardiovascular risk factors, and occupation showed that multivariable-adjusted hazard ratios (HRs) associated with overtime work of ≥11h/day were: 1.63 (95% confidence interval [CI] 1.01-2.63) for acute myocardial infarction and 0.83 (95% CI 0.60-1.13) for total stroke, as compared with the reference group (working 7 to <9 h/day). In the multivariable model, increased risk of acute myocardial infarction associated with overtime work of ≥11 h/day was more evident among salaried employees (HR 2.11, 95% CI 1.03-4.35) and men aged 50-59 years (HR 2.60, 95% CI 1.42-4.77). CONCLUSIONS: Among middle-aged Japanese men, working overtime is associated with a higher risk of acute myocardial infarction.

[Early Liver Metastases after Curative Surgery for Stageâ Gastric Cancer with Neuroendocrine Differentiation-A Case Report].

An 87-year-old male patient was admitted to our hospital with a chief complaint of vomiting. Gastroscopy revealed Type 0-Ⅱc+Ⅱa tumor at the posterior wall in the middle third of the stomach. A biopsy indicated moderately differentiated adenocarcinoma. Abdominal CT revealed no lymph node or distant metastases. The clinical diagnosis was cT2(MP), N0, M0, cStage Ⅰ. Laparoscopic distal gastrectomy with D2 lymphadenectomy was performed. The pathological findings revealed moderately differentiated adenocarcinoma containing synaptophysin, chromogranin A, and CD56-positive tumor cells. He was then diagnosed with adenocarcinoma with neuroendocrine differentiation. The pathological diagnosis was pT2(MP), pN0, M0, pStage ⅠB. MRI revealed multiple liver metastases 5 months postoperatively. S-1 alone chemotherapy was started, and the patient showed partial response(PR)after 3 courses, according to the Response Evaluation Criteria in Solid Tumor (RECIST).

The 5' untranslated region of the rbp1 mRNA is required for translation of its mRNA under low temperatures in the cyanobacterium Synechococcus elongatus.

The unicellular cyanobacterium Synechococcus elongatus has three RNA-binding protein (Rbp) genes, rbp1, rbp2 and rbp3. The rbp1 gene was upregulated by cold treatment while rbp2 and rbp3 expression decreased remarkably after exposure to cold temperatures. To investigate the mechanism underlying cold-induced rbp1 expression, a series of rbp1-luxAB transcriptional fusion constructs were expressed in S. elongatus PCC 7942 under cold conditions. The results showed that the region from -33 to -3 of the transcription initiation site contains an essential sequence for basal transcription of the rbp1 gene and that the 120-bp region (-34 to -153) does not contain critical cis-elements required for cold-shock induction. In contrast, mutational analysis carrying the 5'-untranslated region (UTR) of rbp1-luxAB translational fusions indicated that the 5'-UTR of rbp1 plays an important role in cold induction of the rbp1 gene product. Taken together, we conclude that the cold induction of rbp1 may be regulated at a posttranscriptional level rather than at the transcriptional level.

Occupational physical activity in relation to risk of cardiovascular mortality: The Japan Collaborative Cohort Study for Evaluation for Cancer Risk (JACC Study).

We examined the association between patterns of occupational physical activity (OPA) and mortality from cardiovascular disease (CVD) in a Japanese population. A community-based, prospective cohort of 66,161 men and women aged 40-79years without a history of CVD or cancer at baseline (1988-1990) was followed until 2009. OPA was divided into four types: mostly sitting, sitting and standing (sitting/standing), mostly standing, and standing and walking (standing/walking). During follow-up for a median of 19.2years, 3728 deaths from CVD were registered. Compared with mostly sitting OPA, standing/walking OPA was not associated with a reduced risk of CVD mortality for all subjects, but it was associated with a 20% lower risk of CVD mortality among overweight individuals (body mass index ≥25kg/m(2)). Compared with mostly sitting OPA, mostly standing OPA was associated with an approximately 20% higher risk of CVD mortality, especially among overweight individuals or those with lower exercise (<2.5h/week). In conclusion, compared with mostly sitting OPA, standing/walking OPA is associated with lower CVD mortality among overweight individuals, while mostly standing OPA is associated with higher CVD mortality, especially in physically inactive individuals.

Overexpression of the NADP+-specific isocitrate dehydrogenase gene (icdA) in citric acid-producing Aspergillus niger WU-2223L.

In the tricarboxylic acid (TCA) cycle, NADP(+)-specific isocitrate dehydrogenase (NADP(+)-ICDH) catalyzes oxidative decarboxylation of isocitric acid to form α-ketoglutaric acid with NADP(+) as a cofactor. We constructed an NADP(+)-ICDH gene (icdA)-overexpressing strain (OPI-1) using Aspergillus niger WU-2223L as a host and examined the effects of increase in NADP(+)-ICDH activity on citric acid production. Under citric acid-producing conditions with glucose as the carbon source, the amounts of citric acid produced and glucose consumed by OPI-1 for the 12-d cultivation period decreased by 18.7 and 10.5%, respectively, compared with those by WU-2223L. These results indicate that the amount of citric acid produced by A. niger can be altered with the NADP(+)-ICDH activity. Therefore, NADP(+)-ICDH is an important regulator of citric acid production in the TCA cycle of A. niger. Thus, we propose that the icdA gene is a potentially valuable tool for modulating citric acid production by metabolic engineering.

A case of intestinal intussusception with unique hemorrhagic polyps due to AL amyloidosis and excessive anticoagulation.

Most adult intussusceptions are secondary to various pathological conditions that serve as a lead point. Because of their serious nature, intussusceptions often require emergency surgery. We report a surgical case of amyloidosis associated with intussusception, probably due to polypoid protrusions and bleeding tendencies. An 80-year-old man with abdominal pain was suspected of having jejunal intussusception on computed tomography. He had been prescribed warfarin for atrial fibrillation, and excessive anticoagulation was observed with a prolonged prothrombin time/international normalized ratio of 5.44 at presentation. After the excessive anticoagulation was resolved, emergency surgery was performed. The intussuscepted jejunum was resected, and a 7 cm long dark-red pedunculated polyp was identified as the lead point, which was accompanied by multiple small pedunculated polyps. Histopathological examination showed that these were all hemorrhagic polyps. Amyloid depositions were observed in the muscularis mucosae, submucosa, and the walls of the blood vessels. Immunohistochemical analysis revealed immunoglobulin light chain amyloidosis. This case is informative to discuss the clinical sequelae of gastrointestinal amyloid deposition.

Metastases and treatment-resistant lineages in patient-derived cancer cells of colorectal cancer.

Circulating tumor cells (CTCs) play an important role in metastasis and recurrence. However, which cells comprise the complex tumor lineages in recurrence and are key in metastasis are unknown in colorectal cancer (CRC). CRC with high expression of POU5F1 has a poor prognosis with a high incidence of liver metastatic recurrence. We aim to reveal the key cells promoting metastasis and identify treatment-resistant lineages with established EGFP-expressing organoids in two-dimensional culture (2DOs) under the POU5F1 promotor. POU5F1-expressing cells are highly present in relapsed clinical patients' blood as CTCs. Sorted POU5F1-expressing cells from 2DOs have cancer stem cell abilities and abundantly form liver metastases in vivo. Single-cell RNA sequencing of 2DOs identifies heterogeneous populations derived from POU5F1-expressing cells and the Wnt signaling pathway is enriched in POU5F1-expressing cells. Characteristic high expression of CTLA4 is observed in POU5F1-expressing cells and immunocytochemistry confirms the co-expression of POU5F1 and CTLA4. Demethylation in some CpG islands at the transcriptional start sites of POU5F1 and CTLA4 is observed. The Wnt/ß-catenin pathway inhibitor, XAV939, prevents the adhesion and survival of POU5F1-expressing cells in vitro. Early administration of XAV939 also completely inhibits liver metastasis induced by POU5F1-positive cells.

Surgery for Rectal Cancer With Mixed Reality of 3D Model During Operation: A Case Report.

Background/Aim: Recently, robotic surgery for rectal cancer has become a common minimally invasive surgery. In addition, the technology of augmented and mixed reality is applied in various living environments, including medicine. We successfully performed robotic surgery for rectal cancer with three-dimensional (3D) images as mixed reality (MR) using HoloLens2. Case Report: The patient was diagnosed with rectal cancer by colonoscopy and a positron-emission computed-tomography scan, and we performed robot-assisted anterior resection. The operator used HoloLens2 and performed the surgery while visualizing 3D images of pelvic anatomy with the location of the rectal cancer as hologram. The operation was performed completely and safely, and she was discharged 11 days after surgery with no postoperative complications. Conclusion: This case presents the usefulness of a MR system offering organ visualization as hologram during surgery.

Treatment response prediction of neoadjuvant chemotherapy for rectal cancer by deep learning of colonoscopy images.

In current clinical practice, several treatment methods, including neoadjuvant therapy, are being developed to improve overall survival or local recurrence rates for locally advanced rectal cancer. The response to neoadjuvant therapy is usually evaluated using imaging data collected before and after preoperative treatment or postsurgical pathological diagnosis. However, there is a need to accurately predict the response to preoperative treatment before treatment is administered. The present study used a deep learning network to examine colonoscopy images and construct a model to predict the response of rectal cancer to neoadjuvant chemotherapy. A total of 53 patients who underwent preoperative chemotherapy followed by radical resection for advanced rectal cancer at the Osaka University Hospital between January 2011 and August 2019 were retrospectively analyzed. A convolutional neural network model was constructed using 403 images from 43 patients as the learning set. The diagnostic accuracy of the deep learning model was evaluated using 84 images from 10 patients as the validation set. The model demonstrated a sensitivity, specificity, accuracy, positive predictive value and area under the curve of 77.6% (38/49), 62.9% (22/33), 71.4% (60/84), 74.5% (38/51) and 0.713, respectively, in predicting a poor response to neoadjuvant therapy. Overall, deep learning of colonoscopy images may contribute to an accurate prediction of the response of rectal cancer to neoadjuvant chemotherapy.

Clinical Impact of Two-Week Placement of a Subcutaneous Suction Drain in Preventing Incisional Surgical Site Infection in Open Gastrointestinal Surgery with Class 4 Dirty Wound: A Retrospective Study.

Background: Two recent randomized controlled trials demonstrated the beneficial effects of subcutaneous drainage in preventing incisional surgical site infection (SSI) in colorectal surgery. This study aimed to evaluate the efficacy of subcutaneous suction drains (SSDs) compared with primary skin closure (PC) in class 4 dirty wound surgery. Patients and Methods: Eighty-one patients undergoing open gastrointestinal surgery with class 4 dirty wounds were enrolled in this study, 30 of whom underwent SSD insertion, whereas the other 51 were treated with PC. Because several studies have reported that the median onset of the development of incisional SSI was eight to 13 days after surgery, we used a two-week placement of an SSD. Comparison of patients treated with SSD and PC and multivariable analysis were performed to test the ability of SSD in decreasing the SSI rate. Results: No differences were observed between the two groups in terms of gender, body mass index, American Society of Anesthesiology score, steroid use, presence of diabetes mellitus, peri-operative transfusion, and surgery type. Surgical site infection incidence was lower in the SSD group (6.6%; 2/30) than that in the PC group (23.5%; 12/51; p = 0.069). Multivariable analysis revealed that the presence of diabetes mellitus was an important independent risk factor for incisional SSI, and the placement of an SSD has substantial preventive effects on incisional SSI (p = 0.018 and p = 0.014, respectively). Conclusions: This study suggested the potential importance of a two-week placement of an SSD for preventing incisional SSI in class 4 dirty wound surgery.

Rapid genomic profiling of circulating tumor DNA in non-small cell lung cancer using Oncomine Precision Assay with Genexus™ integrated sequencer.

Background: Genomic profiling of tumors from cancer patients facilitates molecular-guided therapy. The turnaround time is one of important issues to deliver results timely for clinical decisions. The Ion Torrent™ Genexus™ Integrated Sequencer automates all next generation sequencing (NGS) workflows and delivers results within a day. Methods: In this study, we conducted a feasibility study to evaluate the detection rate of genomic alterations from cell-free total nucleic acid (cfTNA, containing cfDNA and cfRNA) of 119 non-small cell lung cancer using Oncomine Precision Assay on Genexus™ Integrated Sequencer. Oncomine Precision Assay (OPA) covers actionable mutations, copy number variations and fusion genes and that are applicable for the selection of targeted therapy. cfTNA isolated from plasma (derived from 14 ml of blood) were subjected to the Genexus system for library construction, templating, sequencing, and data analyses. Results: The sequencing resulted in median overall depth of 35,773× and median molecular coverage of 2,192× with cfTNA input ranged from 11 to 36 ng. Among the 119 samples evaluated, we detected at least one genomic alteration in plasma cfTNA of 79 cases (66%). When comparing to standard-of-care testing, the sensitivity and specificity of mutation detection in non-small cell lung cancer related genes using liquid biopsy with Genexus-OPA ranged between 49-67% and 93-100%, respectively. 59% of actionable mutations, which were present in tumor tissues, were detected by the Genexus- Oncomine Precision Assay using plasma cfTNA. Among the 5 mutations detected from liquid biopsy only, three mutations are of level 1 evidence according to OncoKB database, highlighting the clinical utilities of liquid biopsy in addressing tumor heterogeneity. Extrathoracic metastasis and levels of lactate dehydrogenase (LDH), C-reactive protein (CRP) and Carcinoembryonic Antigen (CEA) are found to be associated with increased circulating tumor DNA detection. Conclusions: The Genexus™ Integrated Sequencer system is an automated, accurate NGS system with short turnaround time (TAT) that could assist clinicians to make more timely decision.

Development of 7SK snRNA Mimics That Inhibit HIV Transcription.

The 332-nucleotide small nuclear RNA (snRNA) 7SK is a highly conserved non-coding RNA that regulates transcriptional elongation. By binding with positive transcriptional elongation factor b (P-TEFb) via HEXIM1, 7SK snRNA decreases the kinase activity of P-TEFb and inhibits transcriptional elongation. Additionally, it is reported that 7SK inhibition results in the stimulation of human immunodeficiency virus (HIV)-specific transcription. These reports suggest that 7SK is a naturally occurring functional molecule as negative regulator of P-TEFb and HIV transcription. In this study, we developed functional oligonucleotides that mimic the function of 7SK (7SK mimics) as novel inhibitors of HIV replication. We defined the essential region of 7SK regarding its suppressive effects on transcriptional downregulation using an antisense strategy. Based on the results, we designed 7SK mimics containing the defined region. The inhibitory effects of 7SK mimics on HIV-1 long terminal repeat promoter specific transcription was drastic compared with those of the control mimic molecule. Notably, these effects were found to be more enhanced by co-transfection with Tat-expressing plasmids. From these results, it is indicated that 7SK mimics may have great therapeutic potential for HIV/AIDS treatment.

Clinical significance of clonal hematopoiesis in the interpretation of blood liquid biopsy.

As the use of next-generation sequencing (NGS) for plasma cell-free DNA (cfDNA) continues to expand in clinical settings, accurate identification of circulating tumor DNA mutations is important to validate its use in the clinical management for cancer patients. Here, we aimed to characterize mutations including clonal hematopoiesis (CH)-related mutations in plasma cfDNA and tumor tissues using the same ultradeep NGS assay and evaluate the clinical significance of CH-related mutations on the interpretation of liquid biopsy results. Ultradeep targeted NGS using Oncomine Pan-Cancer Panel was performed on matched surgically resected tumor tissues, peripheral blood cells (PBCs), and 120 plasma cfDNA samples from 38 colorectal cancer patients. The clinical significance of the CH-related mutations in plasma cfDNA was evaluated by longitudinal monitoring of the postoperative plasma samples. Among the 38 patients, 74 nonsynonymous mutations were identified from tumor tissues and 64 mutations from the preoperative plasma samples. Eleven (17%) of the 64 mutations identified in plasma cfDNA were also detected in PBC DNA and were identified to be CH-related mutations. Overall, 11 of 38 (29%) patients in this cohort harbored at least one CH-related mutation in plasma cfDNA. These CH-related mutations were continuously detected in subsequent postoperative plasma samples from three patients which could be misinterpreted as the presence of residual disease or as lack of treatment response. Our results indicated that it is essential to integrate the mutational information of PBCs to differentiate tumor-derived from CH-related mutations in liquid biopsy analysis. This would prevent the misinterpretation of results to avoid misinformed clinical management for cancer patients.

Molecular cloning and characterization of rat brain endothelial cell derived gene-1 (tumor suppressor candidate 5) expressing abundantly in adipose tissues.

We report the cloning and expressional analysis of rat brain endothelial cell derived gene-1 (BEC-1), detected as a gene dominantly expressed in rat brain endothelial cells by the use of suppression subtractive hybridization technique. The complementary deoxyribonucleic acid sequence of BEC-1 messenger ribonucleic acid was completely determined with a full length of 3410 bp. The open reading frame within the sequence consisted of 522 bp, and the predicted protein sequence was 173 amino acid residues. BEC-1 gene was thought to be rat tumor suppressor candidate 5 (TUSC5), since BEC-1 had considerable homology with both mouse TUSC5 and human located at 17-p-13 point three 1 (LOST1) categorized as human TUSC5 (identities of 97% and 85%, respectively), which were recently identified as a novel tumor suppressor gene candidate. Expressional analyses for BEC-1 mRNA with real-time PCR and of BEC-1 protein by Western blotting demonstrated that both were dominantly expressed in the adipose tissues of Sprague-Dawley (SD) rats. We analyzed and compared the differential expressions of BEC-1 (TUSC5) mRNA and protein in fat tissues between obese homozygous (fa/fa) and lean wild-type (+/+) Zucker rats. Both expressions in the epididymal white adipose tissue (WAT) were highest, followed by those in the interscapular brown adipose tissue (BAT), subcutaneous, and mesenteric WATs, respectively. Interestingly, both expressions in epididymal WAT of obese Zucker rats were significantly lower than those in lean rats. Although cold exposure at 4 degrees C for 6 h significantly stimulated uncoupling protein-1 (UCP-1) mRNA expression, it significantly inhibited BEC-1 (TUSC5) mRNA expression in the interscapular BAT. These data indicated that rat BEC-1 (TUSC5) was abundantly expressed in adipose tissues, and that it might be involved in their regulation independently of UCP-1.

Shortened lifespan of nematode Caenorhabditis elegans after prolonged exposure to heavy metals and detergents.

The acute toxicities of heavy metals and detergents have been well examined with respect to several endpoints, such as mortality, for application to toxicity tests for environmental assessments. However, chronic influences of these agents on multicellular organisms still need to be determined. Here we studied long-term effects on the lifespan of a free-living nematode, Caenorhabditis elegans, resulting from prolonged exposure to heavy metals or detergents, as well as short-term inhibitory effects on reproduction and growth. These agents inhibited growth of hatched larvae and reproductive capacity in a concentration-dependent manner. They also effectively shortened the lifespan of the adult nematode over the same concentration range. Since toxic effects on both growth and lifespan were observed over similar concentration ranges, where acute toxicities in various endpoints are detected, the shortening of the lifespan can be used as a new endpoint for the assessment of various ecotoxic agents.

Development of a drug-coated microneedle array and its application for transdermal delivery of interferon alpha.

Interferon alpha (IFNα) is one of the most famous drugs for the treatment of chronic hepatitis C and various types of human malignancy. Protein drugs, including IFNα, are generally administered by subcutaneous or intramuscular injection due to their poor permeability and low stability in the bloodstream or gastrointestinal tract. Therefore, in the present study, novel IFNα-coated polyvinyl alcohol-based microneedle arrays (IFNα-MNs) were fabricated for the transdermal delivery of IFNα without the painful injection. IFNα was rapidly released from MNs in phosphate buffered solution and these MNs presented piercing ability in the rat skin. Slight erythema and irritation were observed when MNs were applied to the rat skin, but these skin damages completely disappeared within 24 h after removing the IFNα-MNs. Furthermore, the pharmacokinetic parameters of IFNα-MNs were similar to those of IFNα subcutaneous administration. Finally, IFNα-MNs showed a significant antitumor effect in tumor bearing mice similar to that of IFNα subcutaneous administration. These results indicate that IFNα-MNs are a useful biomaterial tool for protein drug therapy and can improve the quality of life in patients by avoidance of painful injections.

Efficient approach to 1,2-diazepines via formal diazomethylene insertion into the C-C bond of cyclobutenones.

Efficient monocyclic 1,2-diazepine formation via a tandem electrocyclization reaction of cyclobutenones with lithiodiazoacetate is demonstrated. The reaction proceeds through an oxy anion-accelerated 4π-ring opening of cyclobutene followed by an 8π-ring closure of the resultant oxy anion-substituted diazo-diene under mild conditions to furnish a 1,2-diazepine via formal diazomethylene insertion into the C-C bond of cyclobutenone.

Development of a novel self-dissolving microneedle array of alendronate, a nitrogen-containing bisphosphonate: evaluation of transdermal absorption, safety, and pharmacological effects after application in rats.

Alendronate is a nitrogen-containing bisphosphonate that is widely used for the treatment of osteoporosis. In this study, we developed a novel self-dissolving micron-size needle array (microneedle array) containing alendronate, which was fabricated by micromodeling technologies using hyaluronic acid as a basic material. Micron-scale pores in the skin were seen after the application of the alendronate-loaded microneedle array, verifying establishment of transdermal pathways for alendronate. The absorption of alendronate after the application of alendronate-loaded microneedle array was almost equivalent to that after subcutaneous administration, and the bioavailability of alendronate was approximately 90% in rats. Furthermore, delivery of alendronate via this strategy effectively suppressed the decrease in the width of the growth plate in a rat model of osteoporosis. Although mild cutaneous irritation was observed after the application of the alendronate-loaded microneedle array, it resolved by day 15. These findings indicate that this alendronate-loaded microneedle array is a promising transdermal formulation for the treatment of osteoporosis.

Development of a novel transdermal patch of alendronate, a nitrogen-containing bisphosphonate, for the treatment of osteoporosis.

Bisphosphonates are widely used for the treatment and prevention of bone diseases, including Paget disease, hypercalcemia of malignancy, and postmenopausal osteoporosis. In this study, we developed a novel transdermal patch of alendronate, a nitrogen-containing bisphosphonate, for the treatment of bone diseases. The maximum permeation fluxes of alendronate through rat and human skin after application of this patch were 1.9 and 0.3 µg/cm(2) per hour, respectively. The bioavailability (BA) of alendronate in rats was approximately 8.3% after the application of alendronate patch and approximately 1.7% after oral administration. These results indicated that the transdermal permeation of alendronate using this patch system was sufficient for the treatment of bone diseases. The plasma calcium level was effectively reduced after application of the alendronate patch in 1α-hydroxyvitamin D(3) -induced hypercalcemia model rats. The alendronate patch also effectively suppressed the decrease in bone mass in model rats with osteoporosis. Modest alendronate-induced erythema of rat skin was observed after application of the alendronate patch. Incorporation of butylhydroxytoluene in the alendronate patch almost completely suppressed this alendronate-induced skin damage while maintaining the transdermal permeation and pharmacologic effects of alendronate. These findings indicate that our novel transdermal delivery system for alendronate is a promising approach to improve compliance and quality of life of patients in the treatment of bone diseases.