Culture of piglet intestinal 3D organoids from cryopreserved epithelial crypts and establishment of cell monolayers

International audienceIntestinal organoids are increasingly being used to study the gut epithelium for digestive disease modeling, or to investigate interactions with drugs, nutrients, metabolites, pathogens, and the microbiota. Methods to culture intestinal organoids are now available for multiple species, including pigs, which is a species of major interest both as a farm animal and as a translational model for humans, for example, to study zoonotic diseases. Here, we give an in-depth description of a procedure used to culture pig intestinal 3D organoids from frozen epithelial crypts. The protocol describes how to cryopreserve epithelial crypts from the pig intestine and the subsequent procedures to culture 3D intestinal organoids. The main advantages of this method are (i) the temporal dissociation of the isolation of crypts from the culture of 3D organoids, (ii) the preparation of large stocks of cryopreserved crypts derived from multiple intestinal segments and from several animals at once, and thus (iii) the reduction in the need to sample fresh tissues from living animals. We also detail a protocol to establish cell monolayers derived from 3D organoids to allow access to the apical side of epithelial cells, which is the site of interactions with nutrients, microbes, or drugs. Overall, the protocols described here is a useful resource for studying the pig intestinal epithelium in veterinary and biomedical research

Usefulness of Home Screening for Promoting Awareness of Impaired Glycemic Status and Utilization of Primary Care in a Low Socio-Economic Setting: A Follow-Up Study in Reunion Island

International audienceBackground: Low socioeconomic settings are characterized by high prevalence of diabetes and difficulty in accessing healthcare. In these contexts, proximity health services could improve healthcare access for diabetes prevention. Our primary objective was to evaluate the usefulness of home screening for promoting awareness of impaired glycemic status and utilization of primary care among adults aged 18-79 in a low socioeconomic setting. Methods: This follow-up study was conducted in 2015-2016 in Reunion Island, a French overseas department in the Indian Ocean. Enrollment and screening occurred on the same day at the home of participants (N = 907). Impaired glycemic status was defined as [glycated hemoglobin (HbA1c) ≥5.7%] OR [fasting capillary blood glucose (FCBG) ≥1.10 g/L] OR [HbA1c = 5.5-5.6% and FCBG = 1.00-1.09 g/L]. Medical, socio-cultural, and socioeconomic characteristics were collected via a face-to-face questionnaire. A one-month telephone follow-up survey was conducted to determine whether participants had consulted a general practitioner (GP) for confirmation of screening results. A multinomial polytomous logistic regression model was used to identify factors independently associated with non-use of GP consultation for confirmation of screening results and nonresponse to the telephone follow-up survey. Results: Prevalence of glycemic abnormalities was 46.0% (95% CI = 42.7-49.2%). Among participants with impaired glycemic status (N = 417), 77.7% (95% CI = 73.7-81.7%) consulted a GP for confirmation of screening results, 12.5% (95% CI = 9.3-15.6%) did not, and 9.8% failed to respond to the follow-up survey. Factors independently associated with nonuse of GP consultation for confirmation of screening results were self-reported unwillingness to consult a GP (adjusted odds ratio [OR]: 4.86, 95% CI = 1.70-13.84), usual GP consultation frequency of less than once a year (adjusted OR: 4.13, 95% CI = 1.56-10.97), and age 18-39 years (adjusted OR: 3.09, 95% CI = 1.46-6.57). Conclusion: Home screening for glycemic abnormalities is a useful proximity health service for diabetes prevention in low socioeconomic settings. Further efforts, including health literacy interventions, are needed to increase utilization of primary care

QUALITY OF LIFE TEN YEARS AFTER A CHIKUNGUNYA OUTBREAK : THE QOL-CHIK POPULATION-BASED STUDY ON REUNION ISLAND

67th Annual Meeting of the American-Society-of-Tropical-Medicine-and-Hygiene (ASTHM), New Orleans, LA, OCT 28-NOV 01, 201

QUALITY OF LIFE TEN YEARS AFTER A CHIKUNGUNYA OUTBREAK : THE QOL-CHIK POPULATION-BASED STUDY ON REUNION ISLAND

67th Annual Meeting of the American-Society-of-Tropical-Medicine-and-Hygiene (ASTHM), New Orleans, LA, OCT 28-NOV 01, 201

Perceived morbidity and community burden after a Chikungunya outbreak: the TELECHIK survey, a population-based cohort study

Abstract Background Persistent disabilities are key manifestations of Chikungunya virus (CHIKV) infection, especially incapacitating polyarthralgia and fatigue. So far, little is known about their impact on health status. The present study aimed at describing the burden of CHIKV prolonged or late-onset symptoms on the self-perceived health of La Réunion islanders. Methods At 18 months after an outbreak of Chikungunya virus, we implemented the TELECHIK survey; a retrospective cohort study conducted on a random sample of the representative SEROCHIK population-based survey. A total of 1,094 subjects sampled for CHIKV-specific IgG antibodies in the setting of La Réunion island in the Indian Ocean, between August 2006 and October 2006, were interviewed about current symptoms divided into musculoskeletal/rheumatic, fatigue, cerebral, sensorineural, digestive and dermatological categories. Results At the time of interview, 43% of seropositive (CHIK+) subjects reported musculoskeletal pain (vs 17% of seronegative (CHIK-) subjects, P P = 0.04), 75% cerebral disorders (vs 57%, P P = 0.001), 18% digestive complaints (vs 15%, P = 0.21), and 36% skin involvement (vs 34%, P = 0.20) on average 2 years after infection (range: 15-34 months). After controlling for confounders such as age, gender, body mass index or major comorbidities in different Poisson regression models, 33% of joint pains were attributable to CHIKV, 10% of cerebral disorders and 7.5% of sensorineural impairments, while Chikungunya did not enhance fatigue states, digestive and skin disorders. Conclusions On average, 2 years after infection 43% to 75% of infected people reported prolonged or late-onset symptoms highly attributable to CHIKV. These manifestations carry a significant burden in the community in the fields of rheumatology, neurology and sensorineural health.</p

RESEARCH ARTICLE Open Access

Perceived morbidity and community burden after a Chikungunya outbreak: the TELECHIK survey, a population-based cohort stud

Mortality and severe morbidity of very preterm infants: comparison of two French cohort studies

International audienceBACKGROUND:In Reunion Island, a French overseas department, the burden of preterm birth and perinatal mortality exceed those observed in mainland France, despite similar access to standard perinatal care. The purpose of the study was to compare the outcome of two cohorts of NICU-admitted very preterm infants born between 24 and 31 weeks of gestation (WG): the registry-based OGP (Observatoire de la Grande Prématurité, Reunion Island, 2008-2013) cohort, and the nationwide EPIPAGE-2 (mainland France, 2011) observational cohort.METHODS:The primary outcome was adverse neonatal outcomes defined as a composite indicator of in-hospital mortality or any of three following severe morbidities: bronchopulmonary dysplasia (BPD), necrotising enterocolitis, or severe neurological injury (periventricular leukomalacia or grade III-IV intraventricular haemorrhages). Logistic regression modelling adjusting for confounders was performed.RESULTS:A total of 1272 very preterm infants from the Reunionese OGP cohort and 3669 peers from the mainland EPIPAGE-2 cohort were compared. Adverse neonatal outcomes were more likely observed in the OGP cohort (32.6% versus 26.6%, p <  0.001), as result of both increased in-hospital mortality across all gestational age strata and increased BPD among the survivors of the 29-31 WG stratum. After adjusting for gestational age, gender and multiple perinatal factors, the risk of adverse neonatal outcomes was higher in the OGP cohort than in the EPIPAGE-2 cohort across all gestational age strata.CONCLUSIONS:Despite similar guidelines for standard perinatal care, very preterm infants born in Reunion Island have a higher risk for death or severe morbidity compared with those born in mainland France

Predictors of Chikungunya rheumatism: a prognostic survey ancillary to the TELECHIK cohort study.

International audienceABSTRACT: INTRODUCTION: Long-lasting relapsing or lingering rheumatic musculoskeletal pain (RMSP) is the hallmark of Chikungunya virus (CHIKV) rheumatism (CHIK-R). Little is known on their prognostic factors. The aim of this prognostic study was to search the determinants of lingering or relapsing RMSP indicative of CHIK-R. METHODS: Three hundred and forty six infected adults (age [greater than or equal to] 15 years) having declared RMSP at disease onset were extracted from the TELECHIK cohort study, Reunion island, and analysed using a multinomial logistic regression model. We also searched for the predictors of CHIKV-specific IgG titres, assessed at the time of a serosurvey, using multiple linear regression analysis. RESULTS: Of these, 111 (32.1%) reported relapsing RMSP, 150 (43.3%) lingering RMSP, and 85 (24.6%) had fully recovered (reference group) on average two years after acute infection. In the final model controlling gender, the determinants of relapsing RMSP were the age 45-59 years (adjusted OR: 2.9, 95% CI: 1.0-8.6) or greater or equal than 60 years (adjusted OR: 10.4, 95% CI: 3.5-31.1), severe rheumatic involvement (fever, at least six joints plus four other symptoms) at presentation (adjusted OR: 3.6, 95% CI: 1.5-8.2), and CHIKV-specific IgG titres (adjusted OR: 3.2, 95% CI: 1.8-5.5, per one unit increase). Prognostic factors for lingering RMSP were age 45-59 years (adjusted OR: 6.4, 95% CI: 1.8-22.1) or greater or equal than 60 years (adjusted OR: 22.3, 95% CI: 6.3-78.1), severe initial rheumatic involvement (adjusted OR: 5.5, 95% CI: 2.2-13.8) and CHIKV-specific IgG titres (adjusted OR: 6.2, 95% CI: 2.8-13.2, per one unit increase). CHIKV specific IgG titres were positively correlated with age, female gender and the severity of initial rheumatic symptoms. CONCLUSIONS: Our data support the roles of age, severity at presentation and CHIKV specific IgG titres for predicting CHIK-R. By identifying the prognostic value of the humoral immune response of the host, this work also suggest a significant contribution of the adaptive immune response to the physiopathology of CHIK-R and should help to reconsider the paradigm of this chronic infection primarily shifted towards the involvement of the innate immune response

The phenotype of the gut region is more stably retained than developmental stage in piglet intestinal organoids

International audienceIntestinal organoids are innovative in vitro tools to study the digestive epithelium. The objective of this study was to generate jejunum and colon organoids from suckling and weaned piglets in order to determine the extent to which organoids retain a location-specific and a developmental stage-specific phenotype. Organoids were studied at three time points by gene expression profiling for comparison with the transcriptomic patterns observed in crypts in vivo . In addition, the gut microbiota and the metabolome were analyzed to characterize the luminal environment of epithelial cells at the origin of organoids. The location-specific expression of 60 genes differentially expressed between jejunum and colon crypts from suckling piglets was partially retained (48%) in the derived organoids at all time point. The regional expression of these genes was independent of luminal signals since the major differences in microbiota and metabolome observed in vivo between the jejunum and the colon were not reproduced in vitro . In contrast, the regional expression of other genes was erased in organoids. Moreover, the developmental stage-specific expression of 30 genes differentially expressed between the jejunum crypts of suckling and weaned piglets was not stably retained in the derived organoids. Differentiation of organoids was necessary to observe the regional expression of certain genes while it was not sufficient to reproduce developmental stage-specific expression patterns. In conclusion, piglet intestinal organoids retained a location-specific phenotype while the characteristics of developmental stage were erased in vitro . Reproducing more closely the luminal environment might help to increase the physiological relevance of intestinal organoids