Microbiome as Mediator of Diet on Colorectal Cancer Risk: The Role of Vitamin D,Markers of Inflammation and Adipokines

Obesity and diet are associated with colorectal cancer (CRC) risk, and microbiome could mediate this risk factor. To investigate this interaction, we performed a case–control study (34 CRC cases and 32 controls) and analyzed fecal microbiota composition using 16S rRNA metabarcoding and sub-sequential shotgun analyses of genomic bacterial DNA to evaluate the role of microbiome and diet in CRC etiology, taking into account vitamin D and other risk biomarkers. Dietary habits were evaluated using a short questionnaire. Multivariate methods for data integration and mediation analysis models were used to investigate causal relationships. CRC cases were significantly more often deficient in vitamin D than controls (p = 0.04); FokI and CYP24A1 polymorphism frequency were different between cases and controls (p = 0.03 and p = 0.02, respectively). A diet poor in fatty fish and rich in carbohydrates was found to be significantly associated with CRC risk (p = 0.011). The mediation analysis confirmed the significant role of the microbiome in mediating CRC risk—increasing levels of Bifidobacteria/Escherichia genera ratio, an indicator of “healthy” intestinal microbiome, can overcome the effect of diet on CRC risk (p = 0.03). This study suggests that microbiome mediates the diet effect on CRC risk, and that vitamin D, markers of inflammation, and adipokines are other factors to consider in order to achieve a better knowledge of the whole carcinogenic process.     

Assessing Health-Related Quality of Life in Children with Coeliac Disease: The Italian Version of CDDUX

We aimed to assess Health-Related Quality of Life (HRQoL) of Italian children and their parents with coeliac disease (CD) using the Coeliac Disease Dutch Questionnaire (CDDUX). The CDDUX underwent a cross-cultural adaptation in a multi-step process, according to international guidelines. A total of 224 children aged between 8–18 years and their parents were prospectively recruited. Cronbach α coefficient was determined as a measure of internal consistency of the questionnaire and inter-children/parent reliability by intraclass correlation coefficient. Univariate and bivariate regression models were used to evaluate correlations between clinical variables and children and parents subclasses of CDDUX and overall mean Paediatric Quality of Life Inventory (PedsQL). The Italian CDDUX proved to be valid and reliable, mean CDDUX total score revealing a neutral evaluation of the quality of life in children 52.6 ± 17.2 and parents 49.5 ± 17.9 (p = 0.07) with strong correlation with PedsQL. The only clinical variable which appeared to affect significantly quality of life both in children and parents was the lower age. A comparison with our results showed remarkable differences in the HRQoL of populations of various nationalities. The Italian version of the CDDUX questionnaire is a simple and reliable tool for assessing the HRQoL in children and adolescents with CD.     

Free Fatty Acids Signature in Human Intestinal Disorders: Significant Association between Butyric Acid and Celiac Disease

Altered circulating levels of free fatty acids (FFAs), namely short chain fatty acids (SCFAs), medium chain fatty acids (MCFAs), and long chain fatty acids (LCFAs), are associated with metabolic, gastrointestinal, and malignant diseases. Hence, we compared the serum FFA profile of patients with celiac disease (CD), adenomatous polyposis (AP), and colorectal cancer (CRC) to healthy controls (HC). We enrolled 44 patients (19 CRC, 9 AP, 16 CD) and 16 HC. We performed a quantitative FFA evaluation with the gas chromatography–mass spectrometry method (GC–MS), and we performed Dirichlet-multinomial regression in order to highlight disease-specific FFA signature. HC showed a different composition of FFAs than CRC, AP, and CD patients. Furthermore, the partial least squares discriminant analysis (PLS-DA) confirmed perfect overlap between the CRC and AP patients and separation of HC from the diseased groups. The Dirichlet-multinomial regression identified only strong positive association between CD and butyric acid. Moreover, CD patients showed significant interactions with age, BMI, and gender. In addition, among patients with the same age and BMI, being male compared to being female implies a decrease of the CD effect on the (log) prevalence of butyric acid in FFA composition. Our data support GC–MS as a suitable method for the concurrent analysis of circulating SCFAs, MCFAs, and LCFAs in different gastrointestinal diseases. Furthermore, and notably, we suggest for the first time that butyric acid could represent a potential biomarker for CD screening.     

Genomic Characterization of hlyF-positive Shiga Toxin–Producing Escherichia coli,Italy and the Netherlands, 2000–2019

Shiga toxin–producing Escherichia coli (STEC) O80:H2 has emerged in Europe as a cause of hemolytic uremic syndrome associated with bacteremia. STEC O80:H2 harbors the mosaic plasmid pR444_A, which combines several virulence genes, including hlyF and antimicrobial resistance genes. pR444_A is found in some extraintestinal pathogenic E. coli (ExPEC) strains. We identified and characterized 53 STEC strains with ExPEC-associated virulence genes isolated in Italy and the Netherlands during 2000–2019. The isolates belong to 2 major populations: 1 belongs to sequence type 301 and harbors diverse stx₂ subtypes, the intimin variant eae-ξ, and pO157-like and pR444_A plasmids; 1 consists of strains belonging to various sequence types, some of which lack the pO157 plasmid, the locus of enterocyte effacement, and the antimicrobial resistance–encoding region. Our results showed that STEC strains harboring ExPEC-associated virulence genes can include multiple serotypes and that the pR444_A plasmid can be acquired and mobilized by STEC strains.     

Human liver stem cell-derived extracellular vesicles reduce injury in a model of normothermic machine perfusion of rat livers previously exposed to a prolonged warm ischemia

Livers from donors after circulatory death (DCD) are a promising option to increase the donor pool, but their use is associated with higher complication rate and inferior graft survival. Normothermic machine perfusion (NMP) keeps the graft at 37°C, providing nutrients and oxygen supply. Human liver stem cell-derived extracellular vesicles (HLSC-EVs) are able to reduce liver injury and promote regeneration. We investigated the efficacy of a reconditioning strategy with HLSC-EVs in an experimental model of NMP. Following total hepatectomy, rat livers were divided into 4 groups: (i) healthy livers, (ii) warm ischemic livers (60 min of warm ischemia), (iii) warm ischemic livers treated with 5 × 10⁸ HLSC-EVs/g-liver, and (iv) warm ischemic livers treated with a 25 × 10⁸ HLSC-EVs/g-liver. NMP lasted 6 h and HLSC-EVs (Unicyte AG, Germany) were administered within the first 15 min. Compared to controls, HLSC-EV treatment significantly reduced transaminases release. Moreover, HLSC-EVs enhanced liver metabolism by promoting phosphate utilization and pH self-regulation. As compared to controls, the higher dose of HLSC-EV was associated with significantly higher bile production and lower intrahepatic resistance. Histologically, this group showed reduced necrosis and enhanced proliferation. In conclusion, HLSC-EV treatment during NMP was feasible and effective in reducing injury in a DCD model with prolonged warm ischemia.     

Clonal composition and persistence of antigen-specific circulating T follicular helper cells

T follicular helper (T_FH) cells play an essential role in promoting B cell responses and antibody affinity maturation in germinal centers (GC). A subset of memory CD4⁺ T cells expressing the chemokine receptor CXCR5 has been described in human blood as phenotypically and clonally related to GC T_FH cells. However, the antigen specificity and relationship of these circulating T_FH (cT_FH) cells with other memory CD4⁺ T cells remain poorly defined. Combining antigenic stimulation and T cell receptor (TCR) Vβ sequencing, we found T cells specific to tetanus toxoid (TT), influenza vaccine (Flu), or Candida albicans (C.alb) in both cT_FH and non-cT_FH subsets, although with different frequencies and effector functions. Interestingly, cT_FH and non-cT_FH cells specific for C.alb or TT had a largely overlapping TCR Vβ repertoire while the repertoire of Flu-specific cT_FH and non-cT_FH cells was distinct. Furthermore, Flu-specific but not C.alb-specific PD-1⁺ cT_FH cells had a “GC T_FH-like” phenotype, with overexpression of IL21, CXCL13, and BCL6. Longitudinal analysis of serial blood donations showed that Flu-specific cT_FH and non-cT_FH cells persisted as stable repertoires for years. Collectively, our study provides insights on the relationship of cT_FH with non-cT_FH cells and on the heterogeneity and persistence of antigen-specific human cT_FH cells.     

p15INK4b, p14ARF, and p16INK4a inactivation in sporadic and neurofibromatosis type 1-related malignant peripheral nerve sheath tumors.

PURPOSE: Malignant peripheral nerve sheath tumor (MPNST) can arise sporadically or in association with neurofibromatosis type 1. Deletions at the 9p21 locus have been reported in these tumors. To additionally characterize the status of this chromosomal region, in this study we performed a comprehensive, mostly PCR-based molecular analysis of the three tumor suppressor genes p15(INK4b), p14(ARF) and p16(INK4a) located at the 9p21 locus in 26 cryopreserved MPNSTs. EXPERIMENTAL DESIGN: Fourteen neurofibromatosis type 1-related and 12 sporadic cases were investigated for homozygous deletion coupled with fluorescent in situ hybridization, promoter methylation, and mutational analysis, as well as m-RNA expression. RESULTS: The results showed that an inactivation of one or more genes occurred in 77% of MPNSTs and was mainly achieved through homozygous deletion (46%), which, in turn, encompassed all of the three tandemly linked genes in 83% of the deleted cases. Promoter methylation was at a less extent involved in gene silencing (18%), and no mutations were found. Loss of function at DNA level strongly correlated with loss of mRNA expression accounting for 80% of the cases. Because of the close relationship between p14(ARF) and TP53 and between p15(INK4b)/p16(INK4a) and Rb, these results support a model of a coinactivation of TP53 and Rb pathways in 75% of MPNSTs, with functional consequences on cell growth control and apoptosis. CONCLUSIONS: The inactivation of the 9p21 locus is a frequent and peculiar hallmark of MPNST genetic profile leading also to an impaired apoptosis that could be taken into account in treatment planning of these tumors.     

Significance of low birthweight for gestational age among very preterm infants

Objective To estimate the risk of specific adverse neonatal events resulting from the combined effects of prematurity and low birthweight in very preterm infants (delivered at 24–31 weeks of gestation)
Design A cohort study of specific adverse neonatal events in preterm infants born at between 24 and 31 weeks of gestation.
Setting Pavia, Italy.
Population Two hundred and thirty singleton infants with sonographically confirmed gestational age, delivered at 24 to 31 weeks of gestation.
Methods To evaluate the impact of a lower than expected birthweight on selected neonatal events independently of gestational age, we calculated birthweight standard deviation scores (differences between actual birthweight and fitted birthweight divided by fitted standard deviation) for each week of gestation.
Results After adjustment for gestational age and other confounders, there was a significant linear trend relating a decreasing birthweight SDS to an increased likelihood of neonatal death, intraventricular haemorrhage, severe respiratory distress syndrome, and acidosis. Compared with infants with SDS 0 ( 50th centile of birthweight), infants with birthweight SDS < −1 (< 16th centile) had increased odds for neonatal death [odds ratio (OR) 3.7, 95% confidence interval (CI) 1.42–9.6], grade III-IV intraventricular haemorrhage (OR 17.5, 95% CI 4.04–75.9), and neonatal acidosis (OR 3.22, 95% CI 1.41–7.4). The significance of birthweight SDS as a predictor of neonatal outcome, however, was lower than that of gestational age.
Conclusions A lower than expected birthweight affects the likelihood of several adverse neonatal events in very preterm infants. However, a decreasing birthweight SDS affects neonatal outcome less than decreasing gestation does.     

男性与女性糖尿病患者致死性冠心病的额外危险：37个前瞻性队列研究的汇总分析

目的：估计男性和女性糖尿病患者致死性冠心病的相对危险。设计：前瞻性队列研究的汇总分析。数据来源：自1966年至2005年3月间Embase和Medline确认并发表的研究，加上源自亚太队列研究协作组的研究，采用正文词组与MeSH主题词相结合的策略进行检索。综述方法：对入选研究的要求为报告有或无糖尿病的男性与女性致死性冠心病的相对危险比较的估计值。如果对估计值仅做年龄调整（而其它因素来经调整——泽者注），则将该研究排除在外。结果：总共纳入37项2型糖尿病和致死性冠心病的研究，涉及447064例患者。糖尿病患者的致死性冠心病发生率显著高于非糖尿病患者（5．4％比1．6％）。与无糖尿病者相比，糖尿病患者发生致死性冠心病的总的相对危险，女性为3．50，95％可信区间为2．70～4．53，显著高于男性患者（2．06，95％可信区间1．81～2．34）。在除外8项仅根据年龄调整的研究后，两性之间危险的差异大大减小，但仍然具有极显著统计学意义。汇集29项经多因素调整估计值的研究，总的相对危险比（女性比男性）为1．46（1．14～1．88）。结论：糖尿病相关的致死性冠心病相对危险，女性比男性高出50％。这种额外的冠心病危险可以通过女性糖尿病患者具有更多不利的心血管危险因素特点以及倾向于对男性患者更注重治疗的可能偏差来解释。