Profile of Nagasaki Islands Study (NaIS): A Population-Based Prospective Cohort Study on Multi-disease.

In an aging society, it is important to visualize the conditions of people living with diseases or disabilities, such as frailty and sarcopenia, and determine the environmental and genetic factors underlying such conditions. Atherosclerosis and arterial stiffness are key conditions between these factors and noncommunicable diseases. In 2014, we launched a population-based prospective open-cohort study, the Nagasaki Islands Study (NaIS), which was conducted in Goto City, located in the remote islands of Nagasaki Prefecture, Japan, mostly involving middle-aged and older residents. We conducted our own health checkups along with the annual standardized checkups organized by the municipality; recruited study participants; and started to follow-up with them for vital status (death), migration, and occurrence of diseases such as myocardial infarction, stroke, fracture, and human T-cell leukemia virus type 1 (HTLV-1) -associated uveitis. Our checkups were conducted as baseline surveys in different areas of Goto City during the fiscal years 2014-2016, secondary surveys during 2017-2019, and tertiary surveys since 2021, consisting of medical interviews, physical examinations, blood and urine tests, body composition measurements, osteoporosis screening, arterial stiffness measurements, carotid ultrasonography, and dental examination. A total of 4,957 residents participated in either the baseline or secondary surveys and were followed-up; and 3,594 and 3,364 residents (aged 27-96 and 28-98 years) participated in the baseline and secondary surveys, respectively. In conclusion, the NaIS has been undertaken to reveal the influence of aging and risk factors of noncommunicable diseases and disabilities, with an aim to contribute towards better healthcare in the future.

Association between periodontitis and chronic kidney disease by functional atherosclerosis status among older Japanese individuals: A cross-sectional study.

AIM: This study aimed to clarify the influence of functional atherosclerosis on the association between periodontitis and chronic kidney disease (CKD). MATERIALS AND METHODS: A cross-sectional study of 998 older Japanese individuals aged 60-99 years who participated in an oral health check-up was conducted. Early and advanced periodontitis were defined as periodontal pocket depth of 4.0-5.9 mm and ≥6.0 mm, respectively. Functional atherosclerosis was defined as cardio-ankle vascular index ≥9.0. RESULTS: Of the 998 study participants, 238 (23.8%) had CKD. No significant associations between periodontitis and CKD were observed in participants without functional atherosclerosis. After adjusting for known cardiovascular risk factors, the odds ratio (OR) (95% confidence interval [CI]) was 1.31 (0.81-2.11) for early periodontitis and 0.74 (0.41-1.34) for advanced periodontitis. Significant positive associations were observed for participants with functional atherosclerosis; the adjusted ORs (95% CIs) were 1.76 (1.04-3.01) for early periodontitis and 1.95 (1.05-3.63) for advanced periodontitis. CONCLUSIONS: A significant positive association between periodontitis and CKD was established for older participants with functional atherosclerosis. No significant associations were observed for those without functional atherosclerosis. These results can help clarify the influence of periodontitis on systemic circulation.

Relationship between Tongue Pressure and Salivary Bacteria in the Older Adults Requiring Long-Term Care.

INTRODUCTION: Aspiration pneumonia, an important issue for the older adults, is caused by an increase in pathogenic microorganisms in the saliva, aspiration, and weakened host immunity. Recently, decreased tongue pressure has been reported to be associated with dysphagia. This study aimed to investigate the relationship between decreased tongue pressure and the number of bacteria in the saliva of the older adults requiring long-term care. METHODS: This cross-sectional study involved 95 older adults requiring long-term care in a facility or at home, eating orally, and who could understand the instructions for measuring tongue pressure. Sex, age, slowness, weakness, shrinking, exhaustion, low activity, number of teeth, functional teeth unit, denture use, oral hygiene, tongue coating index, dry mouth, tongue pressure, and number of bacteria in the saliva were examined. Bacterial counts were analyzed by real-time PCR for total bacteria, total streptococci, methicillin-resistant Staphylococcus aureus, Streptococcus pneumoniae, Pseudomonas aeruginosa, Porphyromonas gingivalis, and Candida albicans. RESULTS: Multiple regression analysis showed that poor oral hygiene and decreased tongue pressure were independent risk factors for increased bacteria in the saliva. Decreased tongue pressure is significantly correlated with an increased number of total bacteria and the presence of P. gingivalis. DISCUSSION/CONCLUSION: Our results suggest that tongue pressure not only maintains the swallowing function but also prevents the increase of bacteria in the saliva of older adults requiring long-term care.

Risk factors for developing medication-related osteonecrosis of the jaw when preserving the tooth that can be a source of infection in cancer patients receiving high-dose antiresorptive agents: a retrospective study.

PURPOSE: To retrospectively investigate whether apical lesion, alveolar bone loss, probing pocket depth, or local infectious symptoms were associated with the onset of medication-related osteonecrosis of the jaw (MRONJ) in patients treated with high-dose antiresorptive agents who did not undergo tooth extraction. METHODS: The study included 92 patients receiving high-dose antiresorptive agent therapy who had teeth with apical lesion ≧ 3 mm, alveolar bone loss ≧ 1/2, probing pocket depth ≧ 4 mm, or local infection symptoms such as swelling, pain, and pus discharge, but did not undergo tooth extraction. Univariate and multivariate Cox regression analyses were performed to determine the relationship between each variable and MRONJ onset. RESULTS: MRONJ developed in 15 of 92 patients (35 of 404 teeth) from 74 to 1883 days (median, 383 days) after the first visit. Multiple Cox regression analysis revealed that a lower number of teeth, diabetes, increased leukocyte count, administration of antiresorptive agents for 180 days or more, local infection symptoms, apical lesion ≧ 3 mm, and probing pocket depth ≧ 4 mm were significantly correlated with the development of MRONJ. CONCLUSION: It is recommended that teeth with apical lesion ≧ 3 mm, probing pocket depth ≧ 4 mm, or local infection symptoms are extracted before or as early as possible after beginning of medication in cancer patients receiving high-dose antiresorptive agent therapy to prevent the development of MRONJ.

Sacroiliac innervation.

PURPOSE: To investigate the innervation pattern of the sacroiliac region, especially with regard to the sacroiliac joint (SIJ). Dorsal SIJ innervation was analyzed and described. Our main hypothesis was that nerves reach the SIJ dorsally, passing ligamental compartments, as this would explain dorsal SIJ pain. METHODS: To examine sacroiliac innervation, we followed the nerves in over 50 specimens over several years. Plastinated slices were evaluated, nerves in the region were stained histologically, and the data were summarized as 3D models. RESULTS: The Rami communicans and posterior branches of the spinal nerves and their branches that form a dorsal sacral plexus and communicating branches, together with corresponding vessels, were observed to form neurovascular bundles embedded by tiny fatty connectives in gaps and tunnels. Branches of L5-S1 pass the inner sacroiliac ligaments (the interosseous sacroiliac ligament and axial interosseous ligament). The outer sacroiliac ligaments (posterior sacroiliac ligaments, long posterior sacroiliac ligament, sacrotuberal ligament, thoracolumbar fascia) are passed by the S1-S4 branches. However, although the paths of these nerves are in the direction of the SIJ, they do not reach it. It is possible that impingement of the neurovascular bundles may result in pain. Moreover, the gaps and tunnels connect to the open dorsal SIJ. CONCLUSION: Our findings suggest that Bogduk's term "sacroiliac pain" correlates to "sacroiliac innervation", which consists of "inner-" and "outer sacroiliac ligament innervation", and to ventral "SIJ pain". The watery gaps and tunnels observed could play a significant role in innervation and thus in the origins of SIJ pain. LEVEL OF EVIDENCE: Individual cross-sectional studies with consistently applied reference standard and blinding.

Efficacy of 3% hydrogen peroxide solution in cleaning tongue coating before and after surgery: a randomized phase II study.

BACKGROUND: Increased bacterial presence in the tongue coating and thereby, the saliva, may be a risk factor for postoperative complications such as surgical site infection or postoperative pneumonia after cancer surgery. However, no method for cleaning tongue coating has been established experimentally. The purpose of this study was to verify the effect of brushing with 3% hydrogen peroxide on suppression of the number of bacteria in tongue coating. METHODS: Sixteen patients with gastric cancer or colorectal cancer undergoing surgery were randomly allocated to control and intervention groups. In the control group, the tongue was brushed for 30 s with a water-moistened toothbrush, while in the intervention group, the tongue was brushed for 30 s with a toothbrush moistened with 3% hydrogen peroxide. Bacterial counts on tongue coating were measured before and 30 s after cleaning the tongue coating using the Rapid Oral Bacteria Quantification System. RESULTS: In the control group, the number of bacteria on the tongue did not decrease significantly after tongue cleaning on the day before surgery, but did on the day after surgery. In contrast, in the intervention group, the number of bacteria on the tongue decreased significantly after tongue cleaning both on the day before and the day after surgery. Furthermore, when comparing the control and intervention groups, the intervention group had a greater reduction effect. CONCLUSIONS: Tongue brushing with 3% hydrogen peroxide is a useful method to reduce the number of bacteria on the tongue in patients with gastrointestinal cancer undergoing surgery. Trial registration jRCTs071200020 (July 3, 2020).

Effect of periosteal reaction in medication-related osteonecrosis of the jaw on treatment outcome after surgery.

INTRODUCTION: Surgical treatment in patients with medication-related osteonecrosis of the jaw (MRONJ) is superior to conservative treatment. However, treatment outcome in patients with periosteal reaction (PR) was significantly poorer than that of those without PR. The purpose of this retrospective study was to analyze the pathophysiology and clinical significance of PR in MRONJ. MATERIALS AND METHODS: Out of 181 patients with MRONJ undergoing surgery, 38 patients with PR were enrolled in the study. CT examinations, histological examinations, and bacteriological examinations using real-time polymerase chain reaction were performed, and the relationship among the opted surgical method, CT findings, and treatment outcome was investigated. RESULTS: The pattern of PR was classified into three types: type 1, new bone is formed parallel to the mandible, and no gap was evident between the mandible and new bone; type 2, new bone is formed parallel to the mandible, and a gap was evident between them; type 3, an irregular shape. Histological examinations revealed inflammatory tissue in the area visualized as a gap on CT. Bacteriological examination showed the presence of bacteria in the type 2 or type 3 PR. Complete cure was observed in 21 of 38 (55.3%) patients, which was lower than the cure rate of 73.4% in 143 patients without PR. The cure rate was significantly lower in cases with type 3 PR or with persistent osteolysis. CONCLUSIONS: It seems that complete resection of both osteolytic area and type 3 PR is necessary to obtain complete healing in patients undergoing marginal mandibulectomy.

Periosteal reaction of medication-related osteonecrosis of the jaw (MRONJ): clinical significance and changes during conservative therapy.

PURPOSE: We previously reported that the periosteal reaction (PR) in medication-related osteonecrosis of the jaw (MRONJ) is a poor prognostic factor in surgical cases, but it is not clear how PR changes during conservative therapy. The purpose of this retrospective study was to compare computed tomography (CT) findings at the first visit and during follow-up visits in MRONJ patients subjected to conservative therapy and to investigate factors associated with the exacerbation of PR during conservative therapy. METHODS: Sixteen patients with MRONJ of the lower jaw who underwent conservative therapy and experienced a PR on CT images at the first visit and underwent CT examination again after 6 months or more were enrolled in the study. Clinical features and CT findings (extent of osteolytic lesion, extent of PR, type of PR, and changes during conservative treatment) were investigated. RESULTS: On the second CT scan, the osteolytic lesion improved in 4 patients, had not changed in 5, and deteriorated in 7, whereas the PR improved in 5 patients, had not changed in 4, and deteriorated in 7 patients. PR was significantly deteriorated in patients who continued to receive antiresorptive agents during conservative treatment and in patients with deteriorated osteolytic lesions. CONCLUSION: PR in MRONJ often expands during conservative therapy and the PR type progresses from the attached type to the gap type, and the irregular type, but discontinuation of antiresorptive agent may improve PR as well as osteolytic lesions.

The Relationship between Dental Findings and Tongue Pressure: A Survey of 745 Community-Dwelling Adults and Elderly Persons in Japan.

INTRODUCTION: Reduced tongue pressure is one of the causes of dysphagia. The purpose of this study was to investigate the relationship between dental findings and tongue pressure, and whether prosthetic treatment prevents reduced tongue pressure. METHODS: This was a cross-sectional study. Participants were 745 community-dwelling adults and elderly persons in the Goto Islands in Nagasaki, who underwent a health checkup for residents in 2015 and 2016. Data were collected on gender; age; grip strength; hemoglobin; Creatinine (Cr); glycated hemoglobin (HbA1c); history of stroke; smoking, drinking, exercise, and walking habits; number of teeth; wearing of removable dentures; functional units of natural teeth (n-FTUs), fixed prostheses (nif-FTUs), and removable dentures (t-FTUs); and tongue pressure. The associations between each variable and tongue pressure were examined using multiple regression analysis. Next, those with 3 or fewer n-FTUs were selected, and differences in tongue pressure were compared between those with 3 or fewer nif-FTUs and those with 4 or more nif-FTUs, using a propensity score matching method. RESULTS: Male gender, weak grip strength, low HbA1c, no drinking, and a low number of teeth were independent factors significantly associated with lower tongue pressure. Among participants with 3 or fewer n-FTUs, the 43 with 4 or more nif-FTUs showed significantly higher tongue pressure than the 43 with 3 or fewer nif-FTUs after propensity score matching, although the number of t-FTUs was not associated with tongue pressure. DISCUSSION/CONCLUSION: Tooth loss was significantly associated with lower tongue pressure. It was suggested that fixed prosthesis treatment might prevent the reduction of tongue pressure, but removable dentures did not have such an effect.

An efficient genetic test flow for multiple congenital anomalies and intellectual disability.

BACKGROUND: Genetic testing has enabled the diagnosis of multiple congenital anomalies and/or intellectual disabilities. However, because of the phenotypic variability in these disorders, selection of an appropriate genetic test can be difficult and complex. For clinical examination, particularly in clinical facilities, a simple and standardized system is needed. METHODS: We compared microarray comparative genomic hybridization and clinical exome sequencing with regard to diagnostic yield, cost, and time required to reach a definitive diagnosis. After first performing G-banding for 200 patients with multiple congenital anomalies and/or intellectual disability, as a subsequent genetic test, microarray and clinical exome sequencing were compared with regard to diagnostic yield, cost, and time required. RESULTS: There was no obvious difference in the diagnostic rate between the two methods; however, clinical exome sequencing was superior in terms of cost and time. In addition, clinical exome sequencing could sufficiently identify copy number variants, and even smaller copy number variants could be identified. CONCLUSIONS: Clinical exome sequencing should be implemented earlier as a genetic test for undiagnosed patients with multiple congenital anomalies and/or intellectual disabilities. Our results can be used to establish inspection methods in clinical facilities.

Association between high psychological distress and poor oral health-related quality of life (OHQoL) in Japanese community-dwelling people: the Nagasaki Islands Study.

BACKGROUND: We investigated the association between psychological distress and oral health status/oral health-related quality of life (OHQoL) in Japanese community-dwelling people. METHODS: We conducted a cross-sectional study using data from the Nagasaki Islands Study. A total of 1183 (455 men and 728 women) has been analyzed in this study. Psychological distress was measured using the Kessler Psychological Distress Scale (K6). Oral health status was measured by dental examination. The OHQoL was measured using the General Oral Health Assessment Index (GOHAI). We defined the total score of ≥5 points on the K6 as high psychological distress (high-K6 group). RESULTS: The multiple linear regression analysis to identify the GOHAI showed that gender, K6, the total number of teeth, the number of dental caries, and visiting a dental clinic within the past 6 months significantly associated with the GOHAI. Among all of these variables, high-K6 (≥ 5) was a substantial contributing factor of the GOHAI (ß = - 0.23, 95% Cl - 2.31 to -1.41, p < 0.0001). CONCLUSIONS: It is likely that the individual with high psychological distress was strongly related to poor OHQoL even in the general population.

Exploring disease-specific methylated CpGs in human male genital abnormalities by using methylated-site display-amplified fragment length polymorphism (MSD-AFLP).

The incidence of male reproductive system disorders, especially hypospadias, has been increasing in developed countries since the latter half of the 20th century. Endocrine-disrupting chemicals from the environment are considered to be involved in hypospadias onset through epigenetic alterations. This pilot study aimed to explore disease-specific methylated CpGs in human patient samples using the methylated-site display-amplified fragment length polymorphism (MSD-AFLP) technique developed by our research group [1]. We compared clinical samples from hypospadias and phimosis patients. Foreskin and blood samples were collected from one- to two-year-old patients with hypospadias (N = 3) and phimosis (N = 3) during surgical treatment. MSD-AFLP analysis showed significantly decreased CpG-methylation levels of genes such as MYH11 and increased CpG-methylation levels of genes such as PLA2G15 in hypospadias patients. Hierarchical clustering analysis showed that genes with significantly altered CpG levels were more markedly altered in DNA from blood than from foreskin. Because of the small number of samples, further investigation is necessary to elucidate the association between variations in CpG levels in foreskin and blood DNA and male genital abnormalities. However, our MSD-AFLP method appears to be a useful tool for exploring disease-specific methylated-CpGs in human epidemiological studies.

Association between number of pairs of opposing posterior teeth, metabolic syndrome, and obesity.

We investigated the relationship between the number of pairs of posterior teeth and metabolic syndrome (MetS), abdominal obesity (AO), and obesity, among Japanese adults. In 2005, 2,807 Japanese adults aged 25-74 years participated in the Survey of Dental Diseases and the National Health and Nutrition Survey. Based on the survey data, BMI, AO (JAS) determined by the Japan Atherosclerosis Society, AO (IDF) by the International Diabetes Federation, and MetS were assessed. Total functional tooth units (t-FTUs) were scored with pairs of opposing posterior teeth, including artificial teeth. Subjects were divided into three categories of chewing ability based on the score of t-FTUs: Poor (if score ≤ 9), Good (if score = 10-11), and Complete (if score = 12). The relationships between chewing ability and BMI ≥ 25, BMI ≥ 30, AO (JAS), AO (IDF), and MetS were tested using univariate and multivariate logistic regression analysis. The chewing ability was significantly associated with MetS, AO, and obesity in the univariate and multivariate logistic regression analysis. Adjusted OR of "Poor" compared to "Complete" were 1.51 (95% confidence interval (CI), 1.24-1.84) for BMI ≥ 25, 2.10 (95% CI 1.40-3.14) for BMI ≥ 30, 1.31 (95% CI 1.07-1.61) for AO (JAS), 1.40 (95% CI 1.15-1.70) for AO (IDF), and 1.34 (95% CI 1.04-1.72) for MetS. All were statistically significant. Preventing tooth loss and maintaining pairs of good chewing ability may be important factors in preventing MetS, AO, and obesity.

Third primary branch of the posterior ramus of the spinal nerve at the thoracolumbar region: a cadaveric study.

PURPOSE: Spinal column procedures require an accurate understanding of neural pathways relative to the anatomic structure. Since Bogduk's report in 1982, it has been known that the human lumbar posterior ramus of the spinal nerve (PRSN) comprise not two but three primary branches at least in some lumbar segments. The purpose of the current study was to examine the existence of the three primary branches in the thoracic and lumbar segments. METHODS: In this study, we investigated the anatomy of the human PRSN in the thoracic and lumbar segments. Ventral dissection was performed in eight cadavers to determine the anatomy of the PRSN between T1 and L5. RESULTS: At the distal end of a given PRSN, the PRSN divided into three primary branches-medial, intermediate and lateral-in 196 out of 272 segments in the thoracic and lumbar regions in eight cadavers. The medial branch supplied the spinalis compartment, and reached the skin. The lateral branch supplied the iliocostalis muscle compartment, and reached skin. The intermediate branch supplied the longissimus muscle and the area between the medial and the lateral branch, which was a seemingly shorter branch. CONCLUSION: The triplication of the primary branch of the PRSN is considered not uncommon. The third branch should be recognized in the literature and in textbooks.

Association between human T cell leukemia virus 1 (HTLV-1) infection and advanced periodontitis in relation to hematopoietic activity among elderly participants: a cross-sectional study.

BACKGROUND: We reported that human T cell leukemia virus 1 (HTLV-1) infection is positively associated with atherosclerosis. Recent evidence has revealed a close association of periodontitis with atherosclerosis, endothelial dysfunction, and disruption of the microcirculation. However, the association between HTLV-1 and advanced periodontitis has not been investigated to date. Since hematopoietic activity is closely linked to endothelial maintenance activity and is known to decline with age, we hypothesized that the state of hematopoietic activity influenced the association between HTLV-1 and advanced periodontitis in elderly participants. METHODS: A cross-sectional study was performed including 822 elderly participants aged 60-99 years who participated in a dental health check-up. Advanced periodontitis was defined as a periodontal pocket ≥ 6.0 mm. Participants were classified as having low or high hematopoietic activity according to the median values of reticulocytes. RESULTS: HTLV-1 infection was positively related to advanced periodontitis among participants with lower hematopoietic activity (lower reticulocyte count), but not among participants with higher hematopoietic activity (higher reticulocyte count). The adjusted odds ratio (95% confidence interval) considering potential confounding factors was 1.92 (1.05-3.49) for participants with a lower reticulocyte count and 0.69 (0.35-1.36) for participants with a higher reticulocyte count. CONCLUSIONS: Among elderly participants, the association between HTLV-1 infection and advanced periodontitis is influenced by hematopoietic activity. Since hematopoietic activity is associated with endothelial maintenance, these findings provide an efficient tool for clarifying the underlying mechanism of the progression of periodontitis among elderly participants.

Association between human T cell leukemia virus type-1 (HTLV-1) infection and advanced periodontitis in relation to atherosclerosis among elderly Japanese: a cross-sectional study.

BACKGROUND: Human T cell leukemia virus type-1 (HTLV-1) stimulates inflammation activity. Our previous study revealed a positive association between asymptomatic HTLV-1 infection and advanced periodontitis among elderly Japanese individuals with low levels of hematopoietic activity (reflected by reticulocyte levels). Since low hematopoietic activity has been correlated with low-grade inflammation and low-grade inflammation is associated with atherosclerosis, the status of atherosclerosis could, in turn, determine the nature of this association. METHODS: To this end, a cross-sectional study of 907 elderly Japanese individuals (aged 60-99 years), who had participated in dental health check-up during the period 2016-2018, was conducted. Advanced periodontitis was defined as periodontal pocket ≥ 6.0 mm. RESULTS: Among the study population, 295 (32.5%) were found to have atherosclerosis defined as a carotid intima-media thickness (CIMT) of ≥ 1.1 mm. HTLV-1 infection was positively associated with advanced periodontitis in participants with atherosclerosis, but no significant associations were observed among the participants without atherosclerosis. The known risk factors' (including reticulocyte and CIMT) adjusted odds ratio (OR) and 95% confidence interval (CI) of advanced periodontitis were OR 2.01 and 95% CI 1.06-3.81 for participants with atherosclerosis and OR 0.61 and 95% CI 0.34-1.12 for participants without atherosclerosis. CONCLUSION: This study found a significant association between HTLV-1 infection and advanced periodontitis among elderly Japanese with atherosclerosis. However, this association is absent in individuals without atherosclerosis, suggesting that atherosclerosis might act as a determinant in the association between HTLV-1 infection and advanced periodontitis among elderly Japanese.

Familial total anomalous pulmonary venous return with 15q11.2 (BP1-BP2) microdeletion.

A 15q11.2 microdeletion (BP1-BP2) is associated with congenital heart diseases (CHDs), developmental delay, and epilepsy. This deletion co-occurs with CHD in 20-30% patients, but a familial case of CHD and a 15q11.2 deletion has not been identified. Here we report the first familial (three siblings) case of total anomalous pulmonary venous return associated with 15q11.2 deletion. Array comparative genomic hybridization identified a ~395 kb deletion at 15q11.2 in patient 1. This deletion was confirmed by fluorescence in situ hybridization in patients 1 and 3 and their asymptomatic father. No deleterious mutation was identified by proband-only exome sequencing of patient 1. One healthy sibling and their mother did not carry the deletion. This deletion is often inherited from asymptomatic parents with an estimated low penetrance of 10.4%. Conversely, we observed high penetrance of this deletion, but secondary copy-number variants or pathogenic variants were not detected in this family.

Methylated site display (MSD)-AFLP, a sensitive and affordable method for analysis of CpG methylation profiles.

BACKGROUND: It has been pointed out that environmental factors or chemicals can cause diseases that are developmental in origin. To detect abnormal epigenetic alterations in DNA methylation, convenient and cost-effective methods are required for such research, in which multiple samples are processed simultaneously. We here present methylated site display (MSD), a unique technique for the preparation of DNA libraries. By combining it with amplified fragment length polymorphism (AFLP) analysis, we developed a new method, MSD-AFLP. RESULTS: Methylated site display libraries consist of only DNAs derived from DNA fragments that are CpG methylated at the 5' end in the original genomic DNA sample. To test the effectiveness of this method, CpG methylation levels in liver, kidney, and hippocampal tissues of mice were compared to examine if MSD-AFLP can detect subtle differences in the levels of tissue-specific differentially methylated CpGs. As a result, many CpG sites suspected to be tissue-specific differentially methylated were detected. Nucleotide sequences adjacent to these methyl-CpG sites were identified and we determined the methylation level by methylation-sensitive restriction endonuclease (MSRE)-PCR analysis to confirm the accuracy of AFLP analysis. The differences of the methylation level among tissues were almost identical among these methods. By MSD-AFLP analysis, we detected many CpGs showing less than 5% statistically significant tissue-specific difference and less than 10% degree of variability. Additionally, MSD-AFLP analysis could be used to identify CpG methylation sites in other organisms including humans. CONCLUSION: MSD-AFLP analysis can potentially be used to measure slight changes in CpG methylation level. Regarding the remarkable precision, sensitivity, and throughput of MSD-AFLP analysis studies, this method will be advantageous in a variety of epigenetics-based research.

Microdeletion of 19p13.3 in a girl with Peutz-Jeghers syndrome, intellectual disability, hypotonia, and distinctive features.

Peutz-Jeghers syndrome (PJS) is a rare autosomal dominant disease characterized by gastrointestinal polyposis and mucocutaneous pigmentation. Germline point mutations in the serine/threonine kinase 11 (STK11) have been identified in about 70% of patients with PJS. Only a few large genomic deletions have been identified. We report on a girl with PJS and multiple congenital anomalies. She had intellectual disability, umbilical hernia, bilateral inguinal hernias, scoliosis, and distinct facial appearance including prominent mandible, smooth philtrum, and malformed ears. She developed lip pigmentation at the age of 12 years but had no gastrointestinal polyps. Array comparative genomic hybridization revealed an approximately 610 kb deletion at 19p13.3, encompassing STK11. Together with previous reports, the identification of common clinical features suggests that microdeletion at 19p13.3 encompassing STK11 constitutes a distinctive phenotype.

Delineation of the KIAA2022 mutation phenotype: two patients with X-linked intellectual disability and distinctive features.

Next-generation sequencing has enabled the screening for a causative mutation in X-linked intellectual disability (XLID). We identified KIAA2022 mutations in two unrelated male patients by targeted sequencing. We selected 13 Japanese male patients with severe intellectual disability (ID), including four sibling patients and nine sporadic patients. Two of thirteen had a KIAA2022 mutation. Patient 1 was a 3-year-old boy. He had severe ID with autistic behavior and hypotonia. Patient 2 was a 5-year-old boy. He also had severe ID with autistic behavior, hypotonia, central hypothyroidism, and steroid-dependent nephrotic syndrome. Both patients revealed consistent distinctive features, including upswept hair, narrow forehead, downslanting eyebrows, wide palpebral fissures, long nose, hypoplastic alae nasi, open mouth, and large ears. De novo KIAA2022 mutations (p.Q705X in Patient 1, p.R322X in Patient 2) were detected by targeted sequencing and confirmed by Sanger sequencing. KIAA2022 mutations and alterations have been reported in only four families with nonsyndromic ID and epilepsy. KIAA2022 is highly expressed in the fetal and adult brain and plays a crucial role in neuronal development. These additional patients support the evidence that KIAA2022 is a causative gene for XLID.