Identification of target cells of human papillomavirus 18 using squamocolumnar junction organoids.

Human papillomavirus 18 (HPV18) is a highly malignant HPV genotype among high-risk HPVs, characterized by the difficulty of detecting it in precancerous lesions and its high prevalence in adenocarcinomas. The cellular targets and molecular mechanisms underlying its infection remain unclear. In this study, we aimed to identify the cells targeted by HPV18 and elucidate the molecular mechanisms underlying HPV18 replication. Initially, we established a lentiviral vector (HPV18LCR-GFP vector) containing the HPV18 long control region promoter located upstream of EGFP. Subsequently, HPV18LCR-GFP vectors were transduced into patient-derived squamocolumnar junction organoids, and the presence of GFP-positive cells was evaluated. Single-cell RNA sequencing of GFP-positive and GFP-negative cells was conducted. Differentially expressed gene analysis revealed that 169 and 484 genes were significantly upregulated in GFP-positive and GFP-negative cells, respectively. Pathway analysis showed that pathways associated with cell cycle and viral carcinogenesis were upregulated in GFP-positive cells, whereas keratinization and mitophagy/autophagy-related pathways were upregulated in GFP-negative cells. siRNA-mediated luciferase reporter assay and HPV18 genome replication assay validated that, among the upregulated genes, ADNP, FHL2, and NPM3 were significantly associated with the activation of the HPV18 early promoter and maintenance of the HPV18 genome. Among them, NPM3 showed substantially higher expression in HPV-related cervical adenocarcinomas than in squamous cell carcinomas, and NPM3 knockdown of HPV18-infected cells downregulated stem cell-related genes. Our new experimental model allows us to identify novel genes involved in HPV18 early promoter activities. These molecules might serve as therapeutic targets in HPV18-infected cervical lesions.

Cells with stem-like properties are associated with the development of HPV18-positive cervical cancer.

The cellular origins of cervical cancer and the histological differentiation of human papillomavirus (HPV)-infected cells remain unexplained. To gain new insights into the carcinogenesis and histological differentiation of HPV-associated cervical cancer, we focused on cervical cancer with mixed histological types. We conducted genomic and transcriptomic analyses of cervical cancers with mixed histological types. The commonality of the cellular origins of these cancers was inferred using phylogenetic analysis and by assessing the HPV integration sites. Carcinogenesis was estimated by analyzing human gene expression profiles in different histological types. Among 42 cervical cancers with known HPV types, mixed histological types were detected in four cases, and three of them were HPV18-positive. Phylogenetic analysis of these three cases revealed that the different histological types had a common cell of origin. Moreover, the HPV-derived transcriptome and HPV integration sites were common among different histological types, suggesting that HPV integration could occur before differentiation into each histological type. Human gene expression profiles indicated that HPV18-positive cancer retained immunologically cold components with stem cell properties. Mixed cervical cancer has a common cellular origin among different histological types, and progenitor cells with stem-like properties may be associated with the development of HPV18-positive cervical cancer.

Linking antigen specific T-cell dynamics in a microfluidic chip to single cell transcription patterns.

A new non-invasive screening profile has been realized that can aid in determining T-cell activation state at single-cell level. Production of activated T-cells with good specificity and stable proliferation is greatly beneficial for advancing adoptive immunotherapy as innate immunological cells are not effective in recognizing and eliminating cancer as expected. The screening method is realized by relating intracellular Ca2+ intensity and motility of T-cells interacting with APC (Antigen Presenting Cells) in a microfluidic chip. The system is tested using APC pulsed with OVA257-264 peptide and its modified affinities (N4, Q4, T4 and V4), and the T-cells from OT-1 mice. In addition, single cell RNA sequencing reveals the activation states of the cells and the clusters from the derived profiles can be indicative of the T-cell activation state. The presented system here can be versatile for a comprehensive application to proceed with T-cell-based immunotherapy and screen the antigen-specific T-cells with excellent efficiency and high proliferation.

Influence of education and subjective financial status on dietary habits among young, middle-aged, and older adults in Japan: a cross-sectional study.

BACKGROUND: Research has suggested an association between lower socioeconomic status (SES) and unhealthy dietary habits. However, differences in the effects of different SES indicators and age remain unclear. The current study addressed this research gap by investigating the relationship between SES and unhealthy dietary habits, specifically focusing on educational attainment and subjective financial status (SFS) among varied age groups. METHODS: Data were derived from a mail survey of 8,464 people living in a suburb of Tokyo, Japan. Participants were classified into three age groups (20-39 years: young adults; 40-64 years: middle-aged adults; and 65-97 years: older adults). SES was assessed based on individual educational attainment and SFS. Unhealthy dietary habits were defined as skipping breakfast and a low frequency of balanced meal consumption. Participants were asked how often they ate breakfast, and those who did not respond "every day" were categorized as "breakfast skippers." Low frequency of balanced meal consumption was defined as eating a meal that included a staple meal, main dish, and side dishes at least twice a day for less than five days per week. Poisson regression analyses with robust variance adjusted for potential covariates were used to determine the interactive effects of educational attainment and SFS on unhealthy dietary habits. RESULTS: Individuals with lower educational attainment across all age groups skipped breakfast more frequently compared to those with higher educational attainment. For older adults, poor SFS was associated with skipping breakfast. Young adults with poor SFS and middle-aged adults with lower educational attainment tended to eat less balanced meals. In addition, an interaction effect was found in older adults, where those with lower education despite good SFS and those with poor SFS despite higher education were at a greater risk of falling into unhealthy diet. CONCLUSIONS: The findings suggested that different SES indicators affect healthy dietary habits in different generations, and therefore, health policies should consider the potential influence of different SES on promoting healthier dietary habits.

Effects of intergenerational contact on social capital in community-dwelling adults aged 25-84 years: a non-randomized community-based intervention.

BACKGROUND: Accumulating social capital in urban areas is essential to improve community health. Previous studies suggested that intergenerational contact may be effective for enhancing social capital. However, no study has examined the effect of intergenerational contact on social capital through a population-based evaluation. This study aimed to investigate the effects of a community-based intervention to increase the frequency of intergenerational contact on social capital among adults aged 25-84 years. METHODS: This study used a non-randomized controlled trial design to conduct a community-based intervention (from March 2016 to March 2019). The study area was Tama ward, Kawasaki city, Kanagawa, Japan. The area comprises five districts; one district was assigned as the intervention group and the other four districts as the control group. We provided the intervention to residents in the intervention group. The intervention comprised three phases: Phase 1 was the preparation term (organizing the project committee); Phase 2 was the implementation term (trained volunteer staff members, conducted the intergenerational greeting campaign, and held intergenerational contact events); and Phase 3 was the transition term (surrendering the lead role of the project to the city hall field workers). In the control group, field workers provided public health services as usual. We conducted mail surveys in September 2016 and November 2018 to assess the effects of the intervention on social capital during Phase 2. Eligible participants were randomly selected from community-dwelling adults aged 25-84 years according to age (10,620 control group individuals and 4479 intervention group individuals). We evaluated social trust, norm of reciprocity, and social support as outcome variables. RESULTS: In total, 2518 participants completed both surveys and were analyzed (control group: 1727; intervention group: 791). We found that social trust (coefficient = 0.065; 95% confidence interval [CI]: 0.006, 0.125) and norm of reciprocity (coefficient = 0.084; 95% CI: 0.020, 0.149) positively changed in the intervention group compared with the control group. CONCLUSIONS: This community-based intervention may contribute to sustaining and improving social capital among community-dwelling adults. TRIAL REGISTRATION: UMIN000046769 (UMIN-CTR); first registered on January 28, 2022 (retrospectively registered).

Examining health risk behaviors of self-employed and employed workers in Japan: a cross-sectional study.

OBJECTIVES: Self-employed workers have a higher risk for adverse health outcomes than employed workers. However, the differences in health risk behaviors by employment status are largely unknown. This study examined differences in health risk behaviors between self-employed and employed (permanent/non-permanent) workers by sex and age (20-59 years, 60-79 years). STUDY DESIGN: This was a cross-sectional study involving community-dwelling adults living in urban cities in Japan. METHODS: In 2019, we conducted a mail survey in Wako city, Saitama, and Fuchu city, Tokyo. In total, 30,315 adults aged ≥18 years were randomly selected, and 14,185 completed the survey (response rate: 46.8%). The participants for analysis were 8538 workers. Health risk behaviors included physical inactivity (<150 min/wk of moderate-to-vigorous physical activity), prolonged sitting (>480 min/d), high-frequency drinking (≥3 d/wk), tobacco use (current smoker), and overweight (body mass index ≥ 25 kg/m2). We also calculated the total number of health risk behaviors. RESULTS: Self-employed workers had more health risk behaviors than permanent and non-permanent employees, with this difference more significant among younger males. In younger males, compared with self-employment, permanent employment was associated with less tobacco use, and non-permanent employment was associated with less physical inactivity, prolonged sitting, high-frequency drinking, and overweight. In younger females, non-permanent employment was associated with less prolonged sitting and overweight than self-employment. In older males and females, the prevalence of physical inactivity was lower in non-permanent employed than in self-employed workers. CONCLUSIONS: Promoting health behaviors among self-employed may be beneficial for reducing health inequalities between self-employed and employed workers.

[A Case of Diffuse Large B Cell Lymphoma of the Extrahepatic Bile Duct Cholangiocarcinoma].

The patient was an 81-year-old man who was hospitalized with poor appetite and obstructive jaundice. An abdominal CT scan showed remarkable thickening of the wall from the cystic duct to extrahepatic bile duct. Endoscopic retrograde cholangiopancreatography( ERCP)revealed stricture at the extrahepatic bile duct. Cholangiocarcinoma was diagnosed and pancreaticoduodenectomy was performed. The histopathological diagnosis was diffuse large B cell lymphoma (DLBCL). The patient was stable after the operation. We present a case report describing the resection of DLBCL of the extrahepatic bile duct along with a review of the literature.

[Information tools commonly used during a natural disaster among information and communication technology device users].

Objective　During a natural disaster, accessing appropriate information is essential to reduce damage to health. Information and community technology (ICT) devices can help in obtaining information. This study aimed to identify the characteristics of information tools commonly used by ICT device users during a natural disaster, and identify associations between sociodemographic factors and Internet-based information tools.Methods　In 2019, 21,300 adults aged 18 years and above living in Fuchu city, Tokyo, were enrolled in our survey. Participants were asked which ICT devices (computer, smartphone, tablet, or mobile phone) they used. Those who used at least one device were classified as ICT device users. To evaluate the information tools commonly used during a natural disaster, participants were asked to select information resources they would use from a list provided (television, radio, Internet search, alert mail, administrative radio system, local government website, neighborhood, family, and friends). We classified Internet search, alert mail, and local government website as Internet-based tools. A Poisson regression model with robust variance was used to assess sex and age differences among ICT device users and clarify associations between sociodemographic factors and the use of Internet-based information tools.Results　The respondents were 9,201 adults (response rate: 43.2%). Among the ICT device users, more than 95% were below 70 years. Moreover, 66.7% of women and 70.6% of men were 80 years or above. More than 80% preferred television to get information during a natural disaster. Over 70% of women below 60 years and men below 70 years preferred to use an Internet search, compared with 7.8% of women aged 80 years or above. Those who selected Internet search were more likely to be women, have a high household income and high educational attainment, be widowed/divorced/single, and be less likely to live alone or be older adults (especially older women). Participants who selected alert mail were more likely to be women and have high educational attainment, and less likely to be widowed/divorced/single and older adults. Those who selected government websites were more likely to be women, have high educational attainment and be widowed/divorced/single, and less likely to live alone, be widowed/divorced/single, and be older adults (especially older women).Conclusion　There were differences by sex and age in information tools selected for use during a natural disaster among ICT device users. Sociodemographic factors were associated with Internet-based tools, and use differed by type of information tool.

[A Case of Gastric Endocrine Carcinoma].

The patient was an 80-year-old man, hospitalized with poor appetite, light-headedness, and black stool. Esophagogastroduodenoscopy showed an ulcerative lesion in cardia, and the tumor was diagnosed as gastric cancer using the biopsy specimens. The patient underwent a gastrectomy with D1-node dissection. Pathologically, the small tumor cells infiltrated the muscularis propria of the gastric wall, and these tumor cells immunohistochemically showed a positive reaction for synaptophysin. Therefore, the tumor was diagnosed as small cell-neuroendocrine carcinoma of the stomach. Metastasis was not observed in regional lymph nodes, and the TNM classification was defined as pStage ⅠB. After surgery, adjuvant chemotherapy was not performed. The patient is well without recurrence for more than 7 months after the surgery. We experienced and report a case of gastric endocrine cell carcinoma that underwent resection and provide a review of the literature.

[A Case of Disseminated Myelocarcinoma with Bone Metastasis 15 Years after Mastectomy and Relatively Long-Term Survival through Multidisciplinary Treatment].

The patient was a 62-year-old woman who had undergone mastectomy in August 2003 for cancer of the right breast. In addition to radiation therapy in the remaining breast, chemotherapy and endocrine therapy were subsequently performed. The patient had a 10-year recurrence-free postoperative course followed by annual surveillance. Beginning in 2016, an elevation in the serum level of tumor markers was detected; however, no accumulations were found in PET-CT over 2 consecutive years(2016 and 2017). Thereafter, serum levels of tumor markers continued to rise, and a PET-CT in 2018 revealed costal accumulations leading to a diagnosis of late-stage bone metastasis in postoperative year 15. Although hormone therapy was resumed, a continuing rise in the serum level of tumor markers led to a diagnosis of diffuse bone metastasis by MRI in 2019. Treatment with abemaciclib was initiated, and with effective radiotherapy, the PS was maintained at 0-1, serum levels of tumor markers decreased, and the lesions themselves continued at SD. However, in June 2020, multiple liver metastases appeared in an abdominothoracic CT scan. The patient refused chemotherapy; this, a BSC policy was adopted. Acute hemolytic anemia occurred immediately thereafter, and the PS gradually deteriorated. The patient died in September 2020, 17 years and 1 month after surgery.

Co-existence of social isolation and homebound status increase the risk of all-cause mortality.

ABSTRACTBackground:Social isolation and homebound statuses are possible risk factors for increased mortality among older adults. However, no study has addressed the impact of accumulation of these two factors on mortality. The aim of this study was to examine whether such accumulation increased the risk of all-cause mortality. METHODS: The analyzed sample was drawn from a mail survey of 1,023 older adults without instrumental activities of daily living disability. Participants were classified into four groups according to the frequency of both face-to-face and non-face-to-face interactions with others (social isolation and non-social isolation) and the frequency of going outdoors (homebound and non-homebound). Social isolation and homebound statuses were defined as having a social interaction less than once a week and going outdoors either every few days or less, respectively. All-cause mortality information during a six-year follow-up was obtained. RESULTS: In total, 78 (7.6%) participants were both socially isolated and homebound. During the follow-up period, 65 participants died, with an overall mortality rate of 10.6 per 1000 person-years. Cox proportional hazards regression analyses demonstrated that older adults who were socially isolated and homebound showed a significantly higher risk of subsequent all-cause mortality compared with healthy adults who were neither socially isolated nor homebound, independent of potential covariates (aHR, 2.19; 95% CI: 1.04-4.63). CONCLUSION: Our results suggest that the co-existence of social isolation and homebound statuses may synergistically increase risk of mortality. Both active and socially integrated lifestyle in later life might play a major role in maintaining a healthy status.

[A Case of Locally Advanced Breast Cancer Successfully Treated with Epirubicin/Cyclophosphamide].

A 66-year-old woman was referred to our hospital with the chief complaint of a huge exposed left breast mass with bleeding. Triple-negative invasive ductal carcinoma of the breast was diagnosed by core needle biopsy. Computed tomogra- phy showed axillary and infraclavicular lymph node metastases. Epirubicin/cyclophosphamide(EC)therapy was started. We reduced the dose to 80%during courses 2-4. After 4 courses of treatment, CT showed a complete response. We reduced the dose to 50% during courses 5-12 and stopped chemotherapy. Five years have passed without recurrence since the first treatment.

[A Case of Pancreatic Neuroendocrine Carcinoma Presenting Multiple Liver Metastasis as an Initial Manifestation for Which Diagnosis Confirmation Was Difficult].

A 67-year-old woman with a history of esophageal cancer(poorly-differentiated squamous cell carcinoma, pStageⅡ) was diagnosed with 2 liver tumors by regular checkup CT 10 years after her operation. We also observed elevated levels of tumor marker CEA. The tumors were suspected to be metastatic although no primary lesion was identified. We performed partial hepatectomy for diagnostic therapy. The pathological diagnosis was adenocarcinoma suggestive of metastatic tumors but the primary lesion remained unknown. Tumor marker levels were elevated 2 months after the operation and we detected a pancreatic tumor, multiple liver tumors, peritoneal dissemination, and para-aortic lymph node metastasis. Therefore, our clinical diagnosis was multiple metastases with primary pancreatic cancer and chemotherapy was performed. We conducted a thorough review of the diagnostic images and repeated the pathological analysis. Immunobiological staining showed that the tumor cells were positive for neuroendocrine markers such as chromogranin A, CD56, and Ki-67. We eventually diagnosed the liver tumors as metastasis from the pancreatic neuroendocrine carcinoma(Grade 3).

[Community Child-Rearing Support Scale: Applicability across generations and differences in the supportive behaviors among generations].

Objectives　Using the Community Child-Rearing Support Scale (CCRSS), which was developed for older adults, we examined 1) the reliability and validity of the scale for multiple generations, as well as the generational differences in the 2) degree and content of supportive behaviors for child rearing and 3) correlates of the behavior, in conjunction with gender differences.Methods　A mail survey was conducted with residents aged 25-84 years who were randomly selected from two cities in Tokyo and the surrounding areas, and responses were obtained from 8918 residents (response rate: 33%). The respondents were grouped as follows: old-aged (65-84 years), middle-aged (50-64 years), and young-aged (25-49 years). We performed the following analyses by age group and gender: 1) Cronbach's reliability coefficient and a confirmatory factor analysis of the CCRSS, 2) analysis of variance of the total CCRSS scores and scores of the three subscales ("children's security and sound growth," "instrumental support to parents," and "emotional support to parents"), and 3) a multiple regression analysis of the total scores in which various individual characteristics were introduced as explanatory variables.Results　The reliability coefficient was over 0.85 for each age group, and the factor analysis showed good model fitness. Compared to the middle- and young-aged groups, the old-aged group was more likely to provide support for "children's security and sound growth," and equally (for women) or more likely (for men) to provide "emotional support to parents," thereby resulting in the highest total score among the three generations. The mean score of "instrumental support to parents," such as taking care of a child, was the highest for young women, although the score was low overall. Having a child or grandchild under 13 years old and stronger generativity were positively associated with child-rearing support for all age groups; however, the degree of associations varied across the groups. While having a child was strongly associated with support among the young-aged group, the association between support and generativity was stronger for older groups. Moreover, participation in neighborhood associations or volunteer groups and high school or lower education were associated with more support among all age groups, whereas living in rental apartments was associated with less support among the young-aged group.Conclusion　Our findings suggest that the CCRSS can be used for multiple generations and that old people play an important role in child rearing in the community. We found both differences and similarities between generations with respect to the correlates of child-rearing support.

[The relationship between intra- and inter-generational exchange and mental health among young and older adults].

Objective　The purpose of this study was to examine the relationship between intra- and inter-generational exchange and mental health among young adults aged 25-49 years and older adults aged 65-84 years.Methods　In 2016, a community-based, cross-sectional survey was conducted. A total of 3,334 young adults (valid response rate: 24.6%), and 3,116 older adults (valid response rate: 46.0%) completed the survey and were included in the analysis. Their mental health was evaluated using the World Health Organization-Five Well-Being Index (WHO-5), and those who scored under 13 or scored less than one on any item were considered to have poor mental health. To evaluate the intra- or inter-generational exchange, participants were asked about their frequency of interaction with people aged 20-49 years or aged ≥70, excluding family and co-workers. Young adults who interacted with people aged 20-49 years or older adults who interacted with people ≥70 were classified as "having intra-generational exchange," while young adults who interacted with people ≥70 years or older adults who interacted with people aged 20-49 years were classified as "having intergenerational exchange." Therefore, individuals who interacted with both generations were classified as "having multi-generational exchange," and those who did not interact with any generation were categorized as "no exchange." We conducted a logistic regression analysis that included mental health as a dependent variable, intra- and inter-generational exchange as independent variables, and gender, age, educational attainment, marital status, living situation, subjective economic status, social participation, employment, self-rated health, and Instrumental Activity of Daily Living as covariates.Results　Of the 3,334 young adults, 61.5% were mentally healthy, 51.3% had intra-generational exchange, 21.9% had inter-generational exchange, 16.5% had multi-generational exchange, and 42.7% did not have any exchange. Of the 3,116 older adults, 65.8% were mentally healthy, 67.9% had intra-generational exchange, 34.3% had inter-generational exchange, 29.9% had multi-generational exchange, and 21.1% did not have any exchange. A logistic regression analysis revealed that "having intra-generational exchange" and "having intergenerational exchange" were significantly related to better mental health in both young adults (intra-generational; Odds ratios (OR) 1.19, 95% confidence intervals (CI) 1.10-1.30: inter-generational; OR 1.13, 95% CI 1.03-1.25) and older adults (intra-generational; OR 1.15, 95% CI 1.02-1.29: inter-generational; OR 1.46, 95% CI 1.30-1.65). Therefore, "having multi-generational exchange" was more strongly related to better mental health compared with "only having intra-generational exchange."Conclusion　Among young and older adults, intra- and inter-generational exchange were related to better mental health, and multi-generational exchange showed the strongest relationship with better mental health.

[Totally Laparoscopic Stomach-Partitioning Gastrojejunostomy for Gastroduodenal Outlet Obstruction for Elderly Cancer Patients].

We have performed totally laparoscopic stomach-partitioning gastrojejunostomy for gastroduodenal outlet obstruction caused by advanced malignancies in 14 cases. We divided 14 patients into 2 groups according to age at surgery: those<80 years of age and thoseB80 years of age. We compared these 2 groups regarding preoperative status, operative findings, and postoperative course. We found that the stage of cancer in the older group was lower than in the younger group, the number of unresectable factors in the older group were fewer than in the younger group, and the postoperative stay in the older group was shorter than in the younger group. There were no serious postoperative complications in either group and all 14 patients were able to drink or eat the day after surgery. These results suggest that palliative, totally laparoscopic stomachpartitioning gastrojejunostomy may be the best option for elderly cancer patients with gastroduodenal outlet obstruction caused by advanced malignancies, even when their general conditions is poor. We should consider this surgical procedure as soon as possible when the patient decides on palliative treatment.

[A Surgical Case of AFP-Producing Gastric Cancer Discovered by Rupture of Liver Metastatic Lesion].

A 57-year-old man visited our hospital with right hypochondralgia. Abdominal contrast CT showed a 10 cm sized mass in S6-7of the liver and abdominal hemorrhage areas. The tumor showed extrahepatic growth. Blood a-fetoprotein(AFP)was high at 4,447.9ng/mL. Hemostasis was performed through emergency transcatheter arterial embolization. At a later date, upper gastrointestinal endoscopy showed a 20 mm sized type 2 lesion in the gastric corpus. Therefore, we performed distal gastrectomy and right hepatic lobectomy. In histopathological findings, the gastric lesion showed identified as a hepatoid adenocarcinoma, which was positive for AFP protein. The liver lesion was negative for AFP protein, but was similar to hepatoid adenocarcinoma, and no fibrosis was observed in the background liver. Therefore, we diagnosed the tumor as a liver metastasis of AFP-producing gastric cancer. On postoperative day 31, CT showed a metastatic lesion in the S1 of the liver and ascites. Chemotherapy was not successful, and the patient died on postoperative day 75. The resection rate of metastatic liver tumor in AFP-producing gastric cancer is low. This is the second case in Japan, wherein a surgery was performed for a ruptured liver metastatic lesion.

[Long-Term Survivor of a Barrett's Esophageal Adenocarcinoma, Who Underwent Three Times Surgical Resection and MultidisciplinaryTherapyfor Lymph Node Recurrences - Report of a Case].

We report a case of a 42-year-old man who underwent 3 times surgical resection for lymph nodes recurrence and multidisciplinary therapy for Stage IV b Barrett's esophageal adenocarcinoma, and was well 6 years and 3 months after the first resection. The prognosis of the recurrence cases after radical recection of the esophageal cancer is extremely poor. Long-term prognosis may be obtained in few patients, but the cases are squamous cell carcinoma in most of the reported cases. The number of Barrett's esophageal adenocarcinoma patients is increasing, but it is not many. There is little reports, and there is no fixed treatment policy.

Non-biased and efficient global amplification of a single-cell cDNA library.

Analysis of single-cell gene expression promises a more precise understanding of molecular mechanisms of a living system. Most techniques only allow studies of the expressions for limited numbers of gene species. When amplification of cDNA was carried out for analysing more genes, amplification biases were frequently reported. A non-biased and efficient global-amplification method, which uses a single-cell cDNA library immobilized on beads, was developed for analysing entire gene expressions for single cells. Every step in this analysis from reverse transcription to cDNA amplification was optimized. By removing degrading excess primers, the bias due to the digestion of cDNA was prevented. Since the residual reagents, which affect the efficiency of each subsequent reaction, could be removed by washing beads, the conditions for uniform and maximized amplification of cDNAs were achieved. The differences in the amplification rates for randomly selected eight genes were within 1.5-folds, which could be negligible for most of the applications of single-cell analysis. The global amplification gives a large amount of amplified cDNA (>100 µg) from a single cell (2-pg mRNA), and that amount is enough for downstream analysis. The proposed global-amplification method was used to analyse transcript ratios of multiple cDNA targets (from several copies to several thousand copies) quantitatively.

[A Patient with Stage IV Gastric Cancer Who Survived More Than Five Years after Gastrectomy].

The prognosis of patients with Stage IV gastric cancer is generally poor. The 5-year overall survival rate is less than 10%. The patient was a 73-year-old man with Stage IV gastric cancer. Before chemotherapy, peritoneal dissemination was observed using staging laparoscopy. The patient received first-line chemotherapy with TS-1 plus CDDP. Renal function worsened and consequently the therapy was stopped. He received 3 courses of chemotherapy with weekly PTX. The peritoneal dissemination had disappeared by the second staging laparoscopy and he underwent distal gastrectomy. The final diagnosis was pT4a, ly2, v1, pN2(4/16),M0, fStage III B. The patient received adjuvant chemotherapy of TS-1 for 4 years and 8months after gastrectomy. More than 5 year after gastrectomy, the patient is alive without recurrence.