The Japan Public Health Center-based Prospective Study for the Next Generation (JPHC-NEXT): Study Design and Participants

Background: Lifestyle and life-environment factors have undergone drastic changes in Japan over the last few decades. Further, many molecular epidemiologic studies have reported that genetic, epigenetic, and other biomarker information may be useful in predicting individual disease risk.Methods: The Japan Public Health Center-based Prospective Study for the Next Generation (JPHC-NEXT) was launched in 2011 to identify risk factors for lifestyle-related disease, elucidate factors that extend healthy life expectancy, and contribute toward personalized healthcare based on our more than 20 years’ experience with the JPHC Study. From 2011 through 2016, a baseline survey was conducted at 16 municipalities in seven prefectures across the country. A self-administered questionnaire was distributed to all registered residents aged 40–74, which mainly asked about lifestyle factors, such as socio-demographic situation, personal medical history, smoking, alcohol and dietary habits. We obtained informed consent from each participant to participate in this long follow-up study of at least 20 years, including consent to the potential use of their residence registry, medical records, medical fee receipts, care insurance etc., and to the provision of biospecimens (blood and urine), including genomic analysis.Results: As of December 31, 2016, we have established a population-based cohort of 115,385 persons (Response rate 44.1%), among whom 55,278 (47.9% of participants) have provided blood and urine samples. The participation rate was slightly higher among females and in the older age group.Conclusion: We have established a large-scale population-based cohort for next-generation epidemiological study in Japan

Communication Tools Used in Cancer Communication with Children: A Scoping Review

Background: Although communication tools might guide healthcare professionals in communicating with children about cancer, it is unclear what kind of tools are used. This scoping review aimed to map the communication tools used in cancer communication among children with cancer, families, and healthcare professionals. Methods: A comprehensive search using PubMed (including MEDLINE), Embase, CENTRAL, PsycINFO, and CINAHL was conducted on 1 August 2021. We mapped communication tools and their impacts. Results: We included 25 studies (9 experimental studies and 16 feasibility studies) of 29 reports and found 21 communication tools. There was a lack of communication tools that were (1) accessible and validated, (2) designed for healthcare professionals, (3) targeted children, families, and healthcare professionals, and (4) were designed to meet the needs of children and families. Experimental studies showed that the communication tools improved children’s knowledge and psychological outcomes (e.g., health locus of control, quality of life, self-efficacy). Conclusion: We mapped communication tools and identified areas that needed further research, including a lack of tools to guide healthcare professionals and share information with children and families. Further research is needed to develop and evaluate these communication tools. Moreover, it is necessary to investigate how communication tools support children, families, and healthcare professionals

Neuropilin 1 Is Essential for Gastrointestinal Smooth Muscle Contractility and Motility in Aged Mice

<div><p>Background and Aims</p><p>Neuropilin 1 (NRP1) is a non-tyrosine kinase receptor for vascular endothelial growth factor (VEGF) and class 3 semaphorins, playing a role in angiogenesis and neuronal axon guidance, respectively. NRP1 is expressed in smooth muscle cells (SMC) but the functional role of NRP1 in SMC has not been elucidated. We therefore investigated the biological relevance of NRP1 in SMC <i>in vivo</i> by generating mice with SMC-specific <i>Nrp1</i> deficiency.</p><p>Methods</p><p>Conditional gene targeting generated SMC-specific <i>Nrp1</i> knockout mice (<i>Nrp1^SMKO</i>) in which Cre recombinase is driven by the smooth muscle-specific myosin heavy chain (<i>smMHC</i>) promoter.</p><p>Results</p><p>SMC-specific <i>Nrp1</i> deficiency resulted in a significant reduction in intestinal length by 6 months, and, by 18 months, in severe constipation, and enlargement of the intestine consistent with chronic intestinal pseudo-obstruction. These effects were associated with significant thinning of the intestinal smooth muscle, and decreased intestinal contractility. Expression of contractile proteins was reduced in <i>Nrp1^SMKO</i> mice, including the smMHC isoform, SMB, whereas we observed a significant increase in the expression of the small-conductance calcium-activated potassium channel 3 (SK3/KCa2.3), implicated in negative regulation of smooth muscle contraction.</p><p>Conclusions</p><p><i>Nrp1</i> deficiency in visceral SMC results in adult-onset defects in gastrointestinal contractility and motility and causes a shift to a less contractile SMC phenotype. These findings indicate a new role for <i>Nrp1</i> in the maintenance of the visceral SMC contractile phenotype required for normal GI motility in aged mice.</p></div

Conditional targeting of <i>Nrp1</i> in SMC.

<p>(A) Schematic diagram of the targeting strategy to generate <i>Nrp1^SMKO</i> mice. The structure of the NRP1 protein shows the major domains in the extracellular (EC), the transmembrane (TM) and cytoplasmic (Cyt) domains. In floxed <i>Nrp1</i> (<i>Nrp1</i>^fl) mice, the exon2 of <i>Nrp1</i> is flanked by two loxP sites, which undergo Cre-mediated recombination in SMC when crossed to the <i>smMHC-Cre</i> mice, resulting in a null <i>Nrp1</i> allele. The positions of primers (F1,F2,R1,R2) used for genotyping are indicated. (B) Breeding scheme used to generate <i>Nrp1^SMKO</i> and heterozygous <i>Nrp1^+/−</i> controls. In the second round of breeding four different genotypes are generated as indicated in parentheses: <i>Nrp1^fl/+</i> (WT), <i>Nrp1^SMHET</i>, <i>Nrp1^+/−</i>, and <i>Nrp1^SMKO</i>. (C) Genotyping of ear notch DNA. A 900bp band indicates the presence of the <i>Cre</i> transgene (top gel). PCR with primers F1 and R1 (middle gel) detects the loxP site and generates 486bp (<i>Nrp1</i> WT) and 700bp (<i>Nrp1</i>^fl, floxed) products. PCR with primers F2 and R2 (bottom gel) detects Cre-mediated recombination of floxed <i>Nrp1</i> and generates 986bp (<i>Nrp1</i> WT), ∼600bp (<i>Nrp1</i>⁻, recombined) and ∼1200bp (<i>Nrp1</i>^fl, unrecombined). (D) GFP fluorescence in <i>smMHC-Cre</i>^+/− (green) shows that Cre recombinase expression is restricted to vascular and visceral SMC. (E) NRP1/SMA immunohistochemistry and X-gal staining of adult colon and thoracic aorta. NRP1 (i,iv) and SMA (ii, v), are expressed in both the circular (CM) and longitudinal (LM) muscle layers, and the muscularis mucosae (arrowhead) of the colon and in the tunica media of the thoracic aorta (arrow). (iii, vi) X-gal staining of sections from a <i>smMHC-Cre/Rosa26R</i> double heterozygote shows that the expression of the lacZ (blue), driven by the smMHC promoter, is restricted to visceral (iii) and vascular (vi) SMC. Boxed regions are magnified views showing expression of NRP1, SMA and LacZ in smooth muscle cells of the colon.</p

Reduced SMC-specific <i>Nrp1</i> expression in <i>Nrp1^SMKO</i>.

<p>(A) qPCR analysis on the whole adult colon: means±SD, n = 3, *<i>P</i><.05, **<i>P</i><.005, ***<i>P</i><.001. (B) Western blots of NRP1 in the isolated smooth muscle layer from the thoracic aorta and in whole colon tissue samples from mice >18 months old. (C) Reduced expression of NRP1 in visceral SMCs is detected by immunohistochemical staining of adult colon tissue sections. Reduced expression, specifically in SMCs, is evident in the <i>Nrp1^SMKO</i> (iii) sample when compared to the <i>Nrp1^fl/+</i> (i) and <i>Nrp1^+/−</i> (ii) samples, and no change in NRP1 expression can be seen in the myenteric plexus of Auerbach (indicated by the arrows).</p

Loss of <i>Nrp1</i> in SMC affects the expression of markers of contractile phenotype.

<p>(A) qPCR analysis of SMC contractile phenotype markers, <i>smMHC</i> (n = 5) and <i>Sm22-alpha</i> (n≥3), on colonic samples from mice ≥16 months old: means±SD; *<i>P</i><.05, **<i>P</i><.005, ***<i>P</i> = .0005. (B) smMHC/SMA immunohistochemistry in colonic tissue from ≥16 months old mice. (C) Semi-quantitative RT-PCR for the SMA and SMB <i>smMHC</i> isoforms in whole colon cDNA from ≥16 month-old mice. Representative results are shown from 3 mice (<i>Nrp1^fl/+</i>, n = 4; <i>Nrp1^+/−</i>, n = 6; <i>Nrp1^SMKO</i>, n = 6; *P = .04). (D) Immunofluorescent staining of SK3 and SMA in colons of ≥16 months old mice shows enhanced expression of SK3 in the colonic SMC (arrows) of <i>Nrp1^SMKO</i> compared to <i>Nrp1^+/−</i> controls. (E) Western blot of SK3 expression in colon lysates from ≥16 month-old mice. Densitometric quantification of SK3 was normalised to beta-actin expression: means±SD; *<i>P</i> = .0233, n = 4.</p

SMC-specific knockout of <i>Nrp1</i> leads to defects in smooth muscle morphology.

<p>(A) SMA immunohistochemistry on cross-sections of adult colon (≥18 months). Arrow points to the muscularis externa, which was significantly thinner in <i>Nrp1^SMKO</i> (*<i>P</i><0.05, <i>Nrp1^fl/+</i> n = 4; <i>Nrp1^+/−</i> n = 5; <i>Nrp1^SMKO</i> n = 5). The lumen size (asterisk) was significantly larger in the <i>Nrp1^SMKO</i>: means±SD, *<i>P</i><0.05, <i>Nrp1^fl/+</i> n = 3; <i>Nrp1^+/−</i> n = 4; <i>Nrp1^SMKO</i> n = 4. (B) SMA immunofluorescent staining of colonic tissue sections.</p

SMC-specific knockout of <i>Nrp1</i> causes severe constipation and GI tract malformations.

<p>(A) Overnight stool counts from ≥18 month-old mice. Mice were housed in a clean cage 24 hours before harvesting the stools. Differences between <i>Nrp1^SMKO</i> and the controls were significant (***<i>P</i><.001, n≥3). (B) Stool size measurement from 8–12 month- and 18–22 months-old mice. Each dot represents an average of six stools collected from one animal. (****<i>P</i><.0001, ***<i>P</i><.001, *<i>P</i> = .01). (C) Bladder length comparison between ≥18 month-old <i>Nrp1^SMKO</i> and the controls, *<i>P</i> = .0217, n≥6. (D) Macroscopic images of dissected <i>Nrp1^fl/+</i>, <i>Nrp1^+/−</i> and <i>Nrp1^SMKO</i>(≥18 months). The colon (Col) and the bladder (Bl) were severely dilated in <i>Nrp1^SMKO</i>. (E) The entire GI tract was dissected from ≥20 months-old <i>Nrp1^fl/+</i>, <i>Nrp1^+/−</i> and <i>Nrp1^SMKO</i> mice. The arrows point to well-pelleted faeces in the colon of the <i>Nrp1^fl/+</i> and <i>Nrp1^+/−</i> controls, while <i>Nrp1^SMKO</i> displayed an impacted colon and caecum (Ce). The length of the colon and small intestine was significantly shorter in the <i>Nrp1^SMKO</i> than the controls (**<i>P</i><.01, n≥3). The width of the proximal colon was also significantly larger in the <i>Nrp1^SMKO</i> compared to the controls (***<i>P</i><.0001, n≥3).</p

Decreased colonic contractility in <i>Nrp1^SMKO</i>.

<p>Organ bath experiments showing the colonic contractile response to increasing doses of KCl (A) and CCh (B) in <i>Nrp1^SMKO</i> and <i>Nrp1^+/−</i> mice (means±SEM). A significant difference in the contractile tension in response to KCl was observed at 80mM (A, *<i>P</i> = .0359, n = 3). (B) Colonic segments of <i>Nrp1^SMKO</i> had a significantly reduced contractility in response to CCh at 3μM (*<i>P</i><.05, **<i>P</i> = .0088, n = 4). (C) Representative spontaneous contractions are shown for colonic segments from three different <i>Nrp1^SMKO</i> and <i>Nrp1^+/−</i> littermates.</p

SMC proliferation and expression of SMEMB.

<p>(A) BrdU staining of colonic tissue sections at P4 (means±SD, n≥3). (B) Analysis of proliferation at P7 revealed a significant increase in SMC proliferation in <i>Nrp1^SMKO</i> compared to the littermate controls (means±SD, *<i>P</i> = .0146, n = 3). (C,D) Western blots of SMEMB in bladder tissue extracts from P7 pups (C) and colonic extracts from adult (≥16 months, D) mice (n≥3).</p