Vaccination With Mouse Dendritic Cells Loaded With an IpaD-IpaB Fusion Provides Protection Against Shigellosis

This work is licensed under a Creative Commons Attribution 4.0 International License.Diarrheal diseases are a major cause of morbidity and mortality worldwide. They are most prevalent in settings with inadequate sanitation, poor hygiene and contaminated water. An important diarrheal pathogen in such settings is Shigella. No commercially available vaccine exists against shigellosis and immunity to the pathogen is serotype-restricted. We have previously shown that a polypeptide fusion of the Type Three Secretion Apparatus (T3SA) proteins IpaB and IpaD (named DBF) was efficacious as a vaccine against Shigella. Vaccination using different administration routes indicated that protection conferred by DBF did not fully correlate with antibodies. To define the immune responses involved in protection, we studied cellular responses to intranasal immunization with the DBF and the adjuvant dmLT. We found dendritic cell (DC) activation at the nasal associated lymphoid tissue (NALT). Activation markers CD86 and MHCII significantly increase in cells from immunized mice. Antigen exposure in vitro further confirmed the upregulation of CD80 and CD40 in primary dendritic cells. Animals immunized with antigen-primed dendritic cells were protected against Shigella infection, at levels comparable to the efficacy of immunization with the protein vaccine formulation. Therefore, we show that antigen-primed DCs are enough to provide immunity, and propose a mechanism of protection against Shigella spp. based on DC-mediated antigen presentation to T cells.NIH R01 AI 007337

Broadly Protective Shigella Vaccine Based on Type III Secretion Apparatus Proteins

Shigella spp. are food- and waterborne pathogens that cause severe diarrheal and dysenteric disease associated with high morbidity and mortality. Individuals most often affected are children under 5 years of age in the developing world. The existence of multiple Shigella serotypes and the heterogenic distribution of pathogenic strains, as well as emerging antibiotic resistance, require the development of a broadly protective vaccine. All Shigella spp. utilize a type III secretion system (TTSS) to initiate infection. The type III secretion apparatus (TTSA) is the molecular needle and syringe that form the energized conduit between the bacterial cytoplasm and the host cell to transport effector proteins that manipulate cellular processes to benefit the pathogen. IpaB and IpaD form a tip complex atop the TTSA needle and are required for pathogenesis. Because they are common to all virulent Shigella spp., they are ideal candidate antigens for a subunit-based, broad-spectrum vaccine. We examined the immunogenicity and protective efficacy of IpaB and IpaD, alone or combined, coadministered with a double mutant heat-labile toxin (dmLT) from Escherichia coli, used as a mucosal adjuvant, in a mouse model of intranasal immunization and pulmonary challenge. Robust systemic and mucosal antibody- and T cell-mediated immunities were induced against both proteins, particularly IpaB. Mice immunized in the presence of dmLT with IpaB alone or IpaB combined with IpaD were fully protected against lethal pulmonary infection with Shigella flexneri and Shigella sonnei. We provide the first demonstration that the Shigella TTSAs IpaB and IpaD are promising antigens for the development of a cross-protective Shigella vaccine

Characterization of a Novel Fusion Protein from IpaB and IpaD of Shigella spp. and Its Potential as a Pan-Shigella Vaccine

Shigellosis is an important disease in the developing world, where about 90 million people become infected with Shigella spp. each year. We previously demonstrated that the type three secretion apparatus (T3SA) proteins IpaB and IpaD are protective antigens in the mouse lethal pulmonary model. In order to simplify vaccine formulation and process development, we have evaluated a vaccine design that incorporates both of these previously tested Shigella antigens into a single polypeptide chain. To determine if this fusion protein (DB fusion) retains the antigenic and protective capacities of IpaB and IpaD, we immunized mice with the DB fusion and compared the immune response to that elicited by the IpaB/IpaD combination vaccine. Purification of the DB fusion required coexpression with IpgC, the IpaB chaperone, and after purification it maintained the highly α-helical characteristics of IpaB and IpaD. The DB fusion also induced comparable immune responses and retained the ability to protect mice against Shigella flexneri and S. sonnei in the lethal pulmonary challenge. It also offered limited protection against S. dysenteriae challenge. Our results show the feasibility of generating a protective Shigella vaccine comprised of the DB fusion

Impact of detergent on biophysical properties and immune response of the IpaDB fusion protein, a candidate subunit vaccine against Shigella species.

Shigella spp. are causative agents of bacillary dysentery, a human illness with high global morbidity levels, particularly among elderly and infant populations. Shigella infects via the fecal-oral route, and its virulence is dependent upon a type III secretion system (T3SS). Two components of the exposed needle tip complex of the Shigella T3SS, invasion plasmid antigen D (IpaD) and IpaB, have been identified as broadly protective antigens in the mouse lethal pneumonia model. A recombinant fusion protein (DB fusion) was created by joining the coding sequences of IpaD and IpaB. The DB fusion is coexpressed with IpaB's cognate chaperone, IpgC, for proper recombinant expression. The chaperone can then be removed by using the mild detergents octyl oligooxyethelene (OPOE) or N,N-dimethyldodecylamine N-oxide (LDAO). The DB fusion in OPOE or LDAO was used for biophysical characterization and subsequent construction of an empirical phase diagram (EPD). The EPD showed that the DB fusion in OPOE is most stable at neutral pH below 55°C. In contrast, the DB fusion in LDAO exhibited remarkable thermal plasticity, since this detergent prevents the loss of secondary and tertiary structures after thermal unfolding at 90°C, as well as preventing thermally induced aggregation. Moreover, the DB fusion in LDAO induced higher interleukin-17 secretion and provided a higher protective efficacy in a mouse challenge model than did the DB fusion in OPOE. These data indicate that LDAO might introduce plasticity to the protein, promoting thermal resilience and enhanced protective efficacy, which may be important in its use as a subunit vaccine

Isotemporal substitution of inactive time with physical activity and time in bed: cross-sectional associations with cardiometabolic health in the PREDIMEDPlus study

Background: This study explored the association between inactive time and measures of adiposity, clinical parameters, obesity, type 2 diabetes and metabolic syndrome components. It further examined the impact of reallocating inactive time to time in bed, light physical activity (LPA) or moderate-to-vigorous physical activity (MVPA) on cardio-metabolic risk factors, including measures of adiposity and body composition, biochemical parameters and blood pressure in older adults. Methods: This is a cross-sectional analysis of baseline data from 2189 Caucasian men and women (age 55-75 years, BMI 27-40 Kg/m2) from the PREDIMED-Plus study (http://www.predimedplus.com/). All participants had ≥3 components of the metabolic syndrome. Inactive time, physical activity and time in bed were objectively determined using triaxial accelerometers GENEActiv during 7 days (ActivInsights Ltd., Kimbolton, United Kingdom). Multiple adjusted linear and logistic regression models were used. Isotemporal substitution regression modelling was performed to assess the relationship of replacing the amount of time spent in one activity for another, on each outcome, including measures of adiposity and body composition, biochemical parameters and blood pressure in older adults. Results: Inactive time was associated with indicators of obesity and the metabolic syndrome. Reallocating 30 min per day of inactive time to 30 min per day of time in bed was associated with lower BMI, waist circumference and glycated hemoglobin (HbA1c) (all p-values < 0.05). Reallocating 30 min per day of inactive time with 30 min per day of LPA or MVPA was associated with lower BMI, waist circumference, total fat, visceral adipose tissue, HbA1c, glucose, triglycerides, and higher body muscle mass and HDL cholesterol (all p-values < 0.05). Conclusions: Inactive time was associated with a poor cardio-metabolic profile. Isotemporal substitution of inactive time with MVPA and LPA or time in bed could have beneficial impact on cardio-metabolic health

Total and Subtypes of Dietary Fat Intake and Its Association with Components of the Metabolic Syndrome in a Mediterranean Population at High Cardiovascular Risk

Background: The effect of dietary fat intake on the metabolic syndrome (MetS) and in turn on cardiovascular disease (CVD) remains unclear in individuals at high CVD risk. Objective: To assess the association between fat intake and MetS components in an adult Mediterranean population at high CVD risk. Design: Baseline assessment of nutritional adequacy in participants (n = 6560, men and women, 55-75 years old, with overweight/obesity and MetS) in the PREvención con DIeta MEDiterránea (PREDIMED)-Plus randomized trial. Methods: Assessment of fat intake (total fat, monounsatured fatty acids: MUFA, polyunsaturated fatty acids: PUFA, saturated fatty acids: SFA, trans-fatty acids: trans-FA, linoleic acid, α-linolenic acid, and ω-3 FA) using a validated food frequency questionnaire, and diet quality using 17-item Mediterranean dietary questionnaire and fat quality index (FQI). Results: Participants in the highest quintile of total dietary fat intake showed lower intake of energy, carbohydrates, protein and fiber, but higher intake of PUFA, MUFA, SFA, TFA, LA, ALA and ω-3 FA. Differences in MetS components were found according to fat intake. Odds (5th vs. 1st quintile): hyperglycemia: 1.3-1.6 times higher for total fat, MUFA, SFA and ω-3 FA intake; low high-density lipoprotein cholesterol (HDL-c): 1.2 higher for LA; hypertriglyceridemia: 0.7 lower for SFA and ω-3 FA intake. Conclusions: Dietary fats played different role on MetS components of high CVD risk patients. Dietary fat intake was associated with higher risk of hyperglycemia

Longitudinal association of dietary acid load with kidney function decline in an older adult population with metabolic syndrome

Background: Diets high in acid load may contribute to kidney function impairment. This study aimed to investigate the association between dietary acid load and 1-year changes in glomerular filtration rate (eGFR) and urine albumin/creatinine ratio (UACR). Methods: Older adults with overweight/obesity and metabolic syndrome (mean age 65 ± 5 years, 48% women) from the PREDIMED-Plus study who had available data on eGFR (n = 5,874) or UACR (n = 3,639) at baseline and after 1 year of follow-up were included in this prospective analysis. Dietary acid load was estimated as potential renal acid load (PRAL) and net endogenous acid production (NEAP) at baseline from a food frequency questionnaire. Linear and logistic regression models were fitted to evaluate the associations between baseline tertiles of dietary acid load and kidney function outcomes. One year-changes in eGFR and UACR were set as the primary outcomes. We secondarily assessed ≥ 10% eGFR decline or ≥10% UACR increase. Results: After multiple adjustments, individuals in the highest tertile of PRAL or NEAP showed higher one-year changes in eGFR (PRAL, β: –0.64 ml/min/1.73 m2; 95% CI: –1.21 to –0.08 and NEAP, β: –0.56 ml/min/1.73 m2; 95% CI: –1.13 to 0.01) compared to those in the lowest category. No associations with changes in UACR were found. Participants with higher levels of PRAL and NEAP had significantly higher odds of developing ≥10% eGFR decline (PRAL, OR: 1.28; 95% CI: 1.07–1.54 and NEAP, OR: 1.24; 95% CI: 1.03–1.50) and ≥10 % UACR increase (PRAL, OR: 1.23; 95% CI: 1.04–1.46) compared to individuals with lower dietary acid load. Conclusions: Higher PRAL and NEAP were associated with worse kidney function after 1 year of follow-up as measured by eGFR and UACR markers in an older Spanish population with overweight/obesity and metabolic syndrome

Longitudinal association of dietary acid load with kidney function decline in an older adult population with metabolic syndrome

Background: Diets high in acid load may contribute to kidney function impairment. This study aimed to investigate the association between dietary acid load and 1-year changes in glomerular filtration rate (eGFR) and urine albumin/creatinine ratio (UACR). Methods: Older adults with overweight/obesity and metabolic syndrome (mean age 65 ± 5 years, 48% women) from the PREDIMED-Plus study who had available data on eGFR (n = 5,874) or UACR (n = 3,639) at baseline and after 1 year of follow-up were included in this prospective analysis. Dietary acid load was estimated as potential renal acid load (PRAL) and net endogenous acid production (NEAP) at baseline from a food frequency questionnaire. Linear and logistic regression models were fitted to evaluate the associations between baseline tertiles of dietary acid load and kidney function outcomes. One year-changes in eGFR and UACR were set as the primary outcomes. We secondarily assessed ≥ 10% eGFR decline or ≥10% UACR increase. Results: After multiple adjustments, individuals in the highest tertile of PRAL or NEAP showed higher one-year changes in eGFR (PRAL, β: -0.64 ml/min/1.73 m2; 95% CI: -1.21 to -0.08 and NEAP, β: -0.56 ml/min/1.73 m2; 95% CI: -1.13 to 0.01) compared to those in the lowest category. No associations with changes in UACR were found. Participants with higher levels of PRAL and NEAP had significantly higher odds of developing ≥10% eGFR decline (PRAL, OR: 1.28; 95% CI: 1.07-1.54 and NEAP, OR: 1.24; 95% CI: 1.03-1.50) and ≥10 % UACR increase (PRAL, OR: 1.23; 95% CI: 1.04-1.46) compared to individuals with lower dietary acid load. Conclusions: Higher PRAL and NEAP were associated with worse kidney function after 1 year of follow-up as measured by eGFR and UACR markers in an older Spanish population with overweight/obesity and metabolic syndrome. Keywords: albuminuria; chronic kidney disease (CKD); dietary acid load; glomerular filtration rate (GFR); kidney function; net endogenous acid production (NEAP); potential renal acid load (PRAL); renal nutrition

Association between solar insolation and a history of suicide attempts in bipolar I disorder

In many international studies, rates of completed suicide and suicide attempts have a seasonal pattern that peaks in spring or summer. This exploratory study investigated the association between solar insolation and a history of suicide attempt in patients with bipolar I disorder. Solar insolation is the amount of electromagnetic energy from the Sun striking a surface area on Earth. Data were collected previously from 5536 patients with bipolar I disorder at 50 collection sites in 32 countries at a wide range of latitudes in both hemispheres. Suicide related data were available for 3365 patients from 310 onset locations in 51 countries. 1047 (31.1%) had a history of suicide attempt. There was a significant inverse association between a history of suicide attempt and the ratio of mean winter solar insolation/mean summer solar insolation. This ratio is smallest near the poles where the winter insolation is very small compared to the summer insolation. This ratio is largest near the equator where there is relatively little variation in the insolation over the year. Other variables in the model that were positively associated with suicide attempt were being female, a history of alcohol or substance abuse, and being in a younger birth cohort. Living in a country with a state-sponsored religion decreased the association. (All estimated coefficients p <0.01). In summary, living in locations with large changes in solar insolation between winter and summer may be associated with increased suicide attempts in patients with bipolar disorder. Further investigation of the impacts of solar insolation on the course of bipolar disorder is needed.Peer reviewe

Intravenous alteplase for stroke with unknown time of onset guided by advanced imaging: systematic review and meta-analysis of individual patient data

Background: Patients who have had a stroke with unknown time of onset have been previously excluded from thrombolysis. We aimed to establish whether intravenous alteplase is safe and effective in such patients when salvageable tissue has been identified with imaging biomarkers. Methods: We did a systematic review and meta-analysis of individual patient data for trials published before Sept 21, 2020. Randomised trials of intravenous alteplase versus standard of care or placebo in adults with stroke with unknown time of onset with perfusion-diffusion MRI, perfusion CT, or MRI with diffusion weighted imaging-fluid attenuated inversion recovery (DWI-FLAIR) mismatch were eligible. The primary outcome was favourable functional outcome (score of 0–1 on the modified Rankin Scale [mRS]) at 90 days indicating no disability using an unconditional mixed-effect logistic-regression model fitted to estimate the treatment effect. Secondary outcomes were mRS shift towards a better functional outcome and independent outcome (mRS 0–2) at 90 days. Safety outcomes included death, severe disability or death (mRS score 4–6), and symptomatic intracranial haemorrhage. This study is registered with PROSPERO, CRD42020166903. Findings: Of 249 identified abstracts, four trials met our eligibility criteria for inclusion: WAKE-UP, EXTEND, THAWS, and ECASS-4. The four trials provided individual patient data for 843 individuals, of whom 429 (51%) were assigned to alteplase and 414 (49%) to placebo or standard care. A favourable outcome occurred in 199 (47%) of 420 patients with alteplase and in 160 (39%) of 409 patients among controls (adjusted odds ratio [OR] 1·49 [95% CI 1·10–2·03]; p=0·011), with low heterogeneity across studies (I2=27%). Alteplase was associated with a significant shift towards better functional outcome (adjusted common OR 1·38 [95% CI 1·05–1·80]; p=0·019), and a higher odds of independent outcome (adjusted OR 1·50 [1·06–2·12]; p=0·022). In the alteplase group, 90 (21%) patients were severely disabled or died (mRS score 4–6), compared with 102 (25%) patients in the control group (adjusted OR 0·76 [0·52–1·11]; p=0·15). 27 (6%) patients died in the alteplase group and 14 (3%) patients died among controls (adjusted OR 2·06 [1·03–4·09]; p=0·040). The prevalence of symptomatic intracranial haemorrhage was higher in the alteplase group than among controls (11 [3%] vs two [&lt;1%], adjusted OR 5·58 [1·22–25·50]; p=0·024). Interpretation: In patients who have had a stroke with unknown time of onset with a DWI-FLAIR or perfusion mismatch, intravenous alteplase resulted in better functional outcome at 90 days than placebo or standard care. A net benefit was observed for all functional outcomes despite an increased risk of symptomatic intracranial haemorrhage. Although there were more deaths with alteplase than placebo, there were fewer cases of severe disability or death. Funding: None