Electrochemical Aptasensing Platform for the Detection of Retinol Binding Protein-4.

Here, we present the results of our the electrochemical aptasensing strategy for retinol binding protein-4 (RBP-4) detection based on a thiolated aptamer against RBP-4 and 6-mercaptohexanol (MCH) directly immobilized on a gold electrode surface. The most important parameters affecting the magnitude of the analytical signal generated were optimized: (i) the presence of magnesium ions in the immobilization and measurement buffer, (ii) the concentration of aptamer in the immobilization solution and (iii) its folding procedure. In this work, a systematic assessment of the electrochemical parameters related to the optimization of the sensing layer of the aptasensor was carried out (electron transfer coefficients (α), electron transfer rate constants (k0) and surface coverage of the thiolated aptamer probe (ΓApt)). Then, under the optimized conditions, the analytical response towards RBP-4 protein, in the presence of an Fe(CN)63-/4- redox couple in the supporting solution was assessed. The proposed electrochemical strategy allowed for RBP-4 detection in the concentration range between 100 and 1000 ng/mL with a limit of detection equal to 44 ng/mL based on electrochemical impedance spectroscopy (EIS). The specificity studies against other diabetes biomarkers, including vaspin and adiponectin, proved the selectivity of the proposed platform. These preliminary results will be used in the next step to miniaturize and test the sensor in real samples.

Correlations of Serum Retinol-Binding Protein and Stromal Cell-Derived Factor-1 with Renal Function in Patients with Diabetic Kidney Disease.

BACKGROUND: The aim is to investigate the correlations of serum retinol-binding protein (RBP) and stromal cell-derived factor-1 (SDF-1) with renal function in patients with diabetic kidney disease (DKD). METHODS: A total of 438 patients with type 2 diabetes mellitus (T2DM) treated from October 2017 to October 2020 were enrolled in this prospective study and divided into simple T2DM and DKD groups. According to urinary albumin-to-creatinine ratio (UACR), DKD patients were divided into moderate, severe, and nephrotic groups. They were assigned to one of the following categories of estimated glomerular filtration rate (eGFR): G1, G2, G3a, G3b, G4, and G5 stages. The correlations of RBP and SDF-1 with renal function were analyzed. RESULTS: The DKD group had a longer T2DM course and higher RBP, uric acid (UA), blood urea nitrogen (BUN), ß2-microglobulin (ß2-MG), serum creatinine (Scr) levels and UACR, and lower SDF-1 level and eGFR than those of simple T2DM group (p < 0.05). The areas under the receiver operating characteristic curves of RBP and SDF-1 for identifying DKD were 0.903 and 0.868, and the optimal cutoff values were 70.71 mg/L and 5.69 ng/mL, respectively. With increasing urinary albumin and clinical stage, RBP, UA, BUN, ß2-MG and Scr levels and UACR significantly rose, while SDF-1 level and eGFR declined (p < 0.05). In patients with DKD, RBP was correlated positively with UACR, UA, BUN, ß2-MG, and Scr (r = 0.764/0.787/0.693/0.577/0.801, p < 0.0001), and negatively with eGFR (r = -0.782, p < 0.0001). SDF-1 was correlated negatively with UACR, UA, BUN, ß2-MG and Scr (r = -0.744/-0.794/-0.666/-0.605/-0.820, p < 0.0001), and positively with eGFR (r = 0.767, p < 0.0001). The multiple linear regression equation was RBP = 29.852 + 0.007 x UACR + 0.101 x UA + 0.497 x BUN + 0.034 x Scr-0.083 x eGFR (p < 0.001). CONCLUSIONS: RBP and SDF-1 can identify DKD in patients with T2DM, and the degree of renal function damage is correlated positively with RBP and negatively with SDF-1. Elevated levels of UA, BUN, Scr and UACR as well as reduced eGFR are risk factors for evaluating RBP.

Influence of geographical environment on retinol-binding protein reference values in Chinese men.

This study combined geographic factors to predict Chinese healthy male RBP reference values from a geographic perspective, with the aim of exploring the spatial distribution and regional differences in Chinese healthy male Retinol-Binding Protein(RBP) reference values, and then providing a theoretical basis for medical diagnosis of healthy male RBP reference values in different regions of China. Using the actual measured RBP values of 24,502 healthy men in 256 cities in China combined with 16 geographical factors as the base data, the spatial autocorrelation, correlation analysis and support vector machine were used to predict the RBP reference values of healthy men in 2322 cities in China, and to generate a spatial distribution map of the RBP reference values of healthy men in China. It was found that the spatial distribution of healthy male RBP reference values in China showed a trend of gradual increase from the first to the third terrain steps. Combined with the distribution map, it is suggested that the RBP reference values of healthy men in China should be divided into the low value zone of the first-level terrain step (25mg/L~40mg/L), the middle value zone of the second-level terrain step (40mg/L~45mg/L) and the high value zone of the third-level terrain step (45mg/L~52mg/L).

Predictive value of serum retinol binding protein in severity and complications of acute pancreatitis: a retrospective cohort study.

OBJECTIVES: Retinol binding protein (RBP) is associated with an increased risk of insulin resistance, metabolic syndrome, atherosclerosis and hypertension. This study aimed to evaluate serum RBP levels in patients with acute pancreatitis (AP). METHODS: The study included 1,871 AP patients, including 1,411 with mild AP (MAP), 244 with moderately severe AP (MSAP), and 186 with severe AP (SAP). Retrospective analysis was conducted on RBP concentrations and other clinical data of AP patients. RESULTS: AP patients were subgrouped by RBP level into low RBP (LRBP), normal RBP (NRBP), and high RBP (HRBP) groups. The LRBP group showed a significantly higher proportion of SAP patients than NRBP and HRBP groups. Additionally, the LRBP group had the highest BISAP and CTSI scores among the three groups; WBC and CRP levels in the NRBP group were significantly lower than those in the LRBP and HRBP groups. RBP was better at predicting acute necrotic collection (ANC) than other local complications, with an area under the curve (AUC) of 0.821. RBP was also an independent risk factor for acute lung injury (ALI) and ANC in AP patients. The AUC of RBP for predicting ALI was 0.829, with 30.45 mg/L as the optimal cutoff value, and the sensitivity and specificity were 59.70% and 96.50%, respectively. The AUC of RBP for predicting ANC was 0.821, with 28.35 mg/L as the optimal cutoff value, and the sensitivity and specificity were 61.20% and 95.50%, respectively. CONCLUSIONS: Serum RBP had predictive value for AP severity, local and systemic complications.

Vitamin A biomarkers were associated with α(1)-acid glycoprotein and C-reactive protein over the course of a human norovirus challenge infection.

Retinol binding protein (RBP) is used as a proxy for retinol in population-based assessments of vitamin A deficiency (VAD) for cost-effectiveness and feasibility. When the cut-off of < 0·7 µmol/l for retinol is applied to RBP to define VAD, an equivalence of the two biomarkers is assumed. Evidence suggests that the relationship between retinol and RBP is not 1:1, particularly in populations with a high burden of infection or inflammation. The goal of this analysis was to longitudinally evaluate the retinol:RBP ratio over 1 month of follow-up among fifty-two individuals exposed to norovirus (n 26 infected, n 26 uninfected), test whether inflammation (measured as α-1-acid glycoprotein (AGP) and C-reactive protein (CRP)) affects retinol, RBP and the ratio between the two and assess whether adjusting vitamin A biomarkers for AGP or CRP improves the equivalence of retinol and RBP. We found that the median molar ratio between retinol and RBP was the same among infected (0·68) and uninfected (0·68) individuals. AGP was associated with the ratio and RBP individually, controlling for CRP, and CRP was associated with both retinol and RBP individually, controlling for AGP over 1 month of follow-up. Adjusting for inflammation led to a slight increase in the ratio among infected individuals (0·71) but remained significantly different from the expected value of one. These findings highlight the need for updated recommendations from the WHO on a cut-off value for RBP and an appropriate method for measuring and adjusting for inflammation when using RBP in population assessments of VAD.

RBP3-Retinopathy-Inherited High Myopia and Retinal Dystrophy: Genetic Characterization, Natural History, and Deep Phenotyping.

PURPOSE: To examine the genetic and clinical features and the natural history of RBP3-associated retinopathy. DESIGN: Multi-center international, retrospective, case series of adults and children, with moleculraly confirmed RBP3-asociated retinopathy. METHODS: The genetic, clinical, and retinal imaging findings, including optical coherence tomography (OCT) and fundus autofluorescence (FAF), were investigated both cross-sectionally and longitudinally. The results of international standard full-field electroretinography (ERG) and pattern electroretinography (PERG) were reviewed. RESULTS: We ascertained 12 patients (5 female and 7 male) from 10 families (4 patients previously reported). Ten novel disease-causing RBP3 variants were identified. Ten patients were homozygous. The mean age (±SD, range) of the group was 21.4 years (±19.1, 2.9-60.5 years) at baseline evaluation. All 12 patients were highly myopic, with a mean spherical equivalent of -16.0D (range, -7.0D to -33.0D). Visual acuity was not significantly different between eyes, and no significant anisometropia was observed. Mean best-corrected visual acuity (BCVA) was 0.48 logMAR (SD, ±0.29; range, 0.2-1.35 logMAR); at baseline. Eleven patients had longitudinal BCVA assessment, with a mean BCVA of 0.46 logMAR after a mean follow-up of 12.6 years. All patients were symptomatic with reduced VA and myopia by the age of 7 years old. All patients had myopic fundi and features in keeping with high myopia on OCT, including choroidal thinning. The 4 youngest patients had no fundus pigmentary changes, with the rest of the patients presenting with a variable degree of mid-peripheral pigmentation and macular changes. FAF showed variable phenotypes, ranging from areas of increased signal to advanced atrophy in older patients. OCT showed cystoid macular edema at presentation in 3 patients, which persisted during follow-up in 2 patients and resolved to atrophy in the third patient. The ERGs were abnormal in 9 of 9 cases, revealing variable relative involvement of rod and cone photoreceptors with additional milder dysfunction post-phototransduction in some. All but 1 patient had PERG evidence of macular dysfunction, which was severe in most cases. CONCLUSIONS: This study details the clinical and functional phenotype of RBP3-retinopathy in the largest cohort reported to date. RBP3-retinopathy is a disease characterized by early onset, slow progression over decades, and high myopia. The phenotypic spectrum and natural history as described herein has prognostic and counseling implications. RBP3-related disease should be considered in children with high myopia and retinal dystrophy.

The level of serum retinol-binding protein is associated with diabetic mild cognitive impairment.

BACKGROUND: Several studies have shown that retinol-binding protein (RBP) is linked to diabetes and neurodegenerative diseases. However, no studies have elucidated the relationship between RBP and diabetic cognitive disorders. OBJECTIVE: To determine whether the change characteristics of serum RBP are associated with alterations in cognitive functioning in type 2 diabetes mellitus (T2DM). METHODS: In this study, 252 patients with T2DM and 34 people as healthy controls were included. According to the Montreal Cognitive Assessment (MoCA), the diabetic subjects were divided into the mild cognitive impairment (MCI) group and the Non-MCI group. Demographic characteristics and clinical indicators as well as serum RBP levels were analyzed. RESULTS: The serum RBP levels in the MCI group were lower compared with the Non-MCI group (P = 0.02). The level of RBP was higher in the diabetes without MCI group than in the healthy control (P < 0.001). Serum RBP levels were positively correlated with MoCA scores (r = 0.178, P = 0.003). Binary Logistic regression model analysis showed that low RBP [odds ratio (OR) = 0.936], old age (OR = 1.074), high fasting blood glucose (OR = 1.164), and low fasting C-peptide (OR = 0.722) may be independent risk factors for diabetic MCI. The ROC curve of serum RBP for predicting diabetic MCI showed that the area under the curve was 0.630. CONCLUSIONS: Our study revealed an association between serum RBP and diabetic MCI. Serum RBP levels in diabetic MCI are lower and correlated with cognitive function.

Urinary orosomucoid and retinol binding protein levels as early diagnostic markers for diabetic kidney Disease.

BACKGROUND: Diabetic kidney disease (DKD) is the most common microvascular complication of diabetes, which has been a major cause of end-stage renal failure. Diagnosing diabetic kidney disease is important to prevent long-term kidney damage and determine the prognosis of patients with diabetes. In this study, we investigated the clinical significance of combined detection of urine orosomucoid and retinol-binding protein for early diagnosis of diabetic kidney disease. METHODS: We recruited 72 newly diagnosed patients with type 2 diabetes and 34 healthy persons from August 2016 to July 2018 at the First Affiliated Hospital of Henan Polytechnic University (Jiaozuo Second People's Hospital). Using the Mogensen grading criteria, participants were classified as having diabetes or diabetic kidney disease, and healthy persons constituted the control group. Urine orosomucoid and retinol-binding protein levels were measured and correlated with other variables. RESULTS: With the aggravation of renal damage, the level of urinary mucoid protein gradually increased. Urinary retinol-binding protein and microalbumin levels were significantly higher in the diabetes group than in control and nephropathy groups. Orosomucoid and retinol-binding protein might be independent risk factors for diabetes and diabetic kidney disease. Urinary orosomucoid significantly correlated with retinol-binding protein and microalbumin levels in the diabetic kidney disease group. CONCLUSION: Elevated urine orosomucoid and retinol-binding protein levels can be detected in the early stages of type 2 diabetic kidney disease. Both of these markers are important for diabetic kidney disease detection and early treatment.

Significance of Plasma Irisin, Adiponectin, and Retinol Binding Protein-4 Levels as Biomarkers for Obstructive Sleep Apnea Syndrome Severity.

OBJECTIVE: Obstructive sleep apnea syndrome (OSAS) is a common sleep disorder that is caused by the reduction or cessation of airflow in the upper airway. Irisin, retinol-binding protein-4 (RBP-4), and adiponectin are the three significant factors in the metabolic process of the human body. The objective of this study was to investigate whether plasma irisin, RBP-4, and adiponectin levels are associated with the severity of OSAS. METHODS: According to inclusion and exclusion criteria, 125 patients with OSAS and 46 healthy, gender-matched controls were included in this study. The patients were classified according to the apnea hypopnea index (AHI) as 14 mild cases (5 < AHI < 15), 23 moderate OSAS cases (15 < AHI < 30), and 88 severe OSAS cases (AHI > 30). The plasma irisin, RBP-4, and adiponectin levels were measured and compared between groups. RESULTS: RBP-4 levels were higher in severe OSAS compared to other groups, and irisin levels were significantly lower in severe OSAS compared to other groups. There was a negative correlation between irisin and RBP-4 (r = -0.421; p < 0.001), and irisin and AHI (r = -0.834; p < 0.001), and a positive correlation between irisin and adiponectin (r = 0.240; p = 0.002). There was a negative correlation between RBP-4 and adiponectin (r = -0.507; p < 0.001) and a positive correlation between RBP-4 and AHI (r = 0.473; p < 0.001). As a predictor of OSAS, adiponectin showed the highest specificity (84.8%) and RBP-4 the highest sensitivity (92.0%). CONCLUSION: Circulating adiponectin, irisin, and RBP-4 may be new biomarkers in OSAS patients in addition to risk factors such as diabetes, obesity, and hypertension. When polysomnography is not available, these parameters and clinical data can be used to diagnose the disease. As a result, patients with an AHI score greater than thirty should be closely monitored for metabolic abnormalities.

The profile of adipokines associated with fibrosis and impaired microcirculation in systemic sclerosis.

PURPOSE: Adipokines belong to a group of molecules mostly produced by adipose tissue. Abnormalities in the secretion of several adipokines have already implicated to play a pathogenic role in systemic sclerosis (SSc). However, the possible role of numerous molecules still needs to be clarified. The aim of the study was to determine whether the altered level of selected circulating adipokines might correlate with the intensity of fibrosis and vasculopathy in the course of SSc. MATERIALS AND METHODS: Serum concentrations of chemerin, adipsin, retinol-binding protein 4, apelin, visfatin, omentin-1, and vaspin were determined with ELISA in the sera of patients with SSc (n â€‹= â€‹55) and healthy controls (n â€‹= â€‹25). RESULTS: The serum concentration of adipsin (p â€‹= â€‹0.03) and visfatin (p â€‹= â€‹0.04) was significantly increased and the level of retinol-binding protein 4 (p â€‹= â€‹0.03) was decreased in diffuse compared to limited cutaneous SSc. Moreover, serum adipsin level correlated positively with the intensity of skin fibrosis measured with the modified Rodnan skin score (r â€‹= â€‹0.31, p â€‹= â€‹0.02) and was significantly higher in patients with pulmonary arterial hypertension than in those without the condition (p â€‹= â€‹0.03). The concentrations of adipsin (p â€‹= â€‹0.01) and visfatin (p â€‹= â€‹0.04) were significantly increased and the level of apelin (p â€‹= â€‹0.02) was decreased in patients with active digital ulcerations compared to individuals without this complication. CONCLUSION: Adipsin may be considered a pivotal protein in the development of both fibrosis and impaired microcirculation. Its abnormal concentration reflects the intensity of skin thickening and the presence of pulmonary arterial hypertension. Adipsin, visfatin, and apelin are adipose tissue-derived molecules associated with digital vasculopathy.

Clinical, genetic and biochemical signatures of RBP4-related ocular malformations.

BACKGROUND: The retinoic acid (RA) pathway plays a crucial role in both eye morphogenesis and the visual cycle. Individuals with monoallelic and biallelic pathogenic variants in retinol-binding protein 4 (RBP4), encoding a serum retinol-specific transporter, display variable ocular phenotypes. Although few families have been reported worldwide, recessive inherited variants appear to be associated with retinal degeneration, while individuals with dominantly inherited variants manifest ocular development anomalies, mainly microphthalmia, anophthalmia and coloboma (MAC). METHODS: We report here seven new families (13 patients) with isolated and syndromic MAC harbouring heterozygous RBP4 variants, of whom we performed biochemical analyses. RESULTS: For the first time, malformations that overlap the clinical spectrum of vitamin A deficiency are reported, providing a link with other RA disorders. Our data support two distinct phenotypes, depending on the nature and mode of inheritance of the variants: dominantly inherited, almost exclusively missense, associated with ocular malformations, in contrast to recessive, mainly truncating, associated with retinal degeneration. Moreover, we also confirm the skewed inheritance and impact of maternal RBP4 genotypes on phenotypical expression in dominant forms, suggesting that maternal RBP4 genetic status and content of diet during pregnancy may modify MAC occurrence and severity. Furthermore, we demonstrate that retinol-binding protein blood dosage in patients could provide a biological signature crucial for classifying RBP4 variants. Finally, we propose a novel hypothesis to explain the mechanisms underlying the observed genotype-phenotype correlations in RBP4 mutational spectrum. CONCLUSION: Dominant missense variants in RBP4 are associated with MAC of incomplete penetrance with maternal inheritance through a likely dominant-negative mechanism.

Association with controlling nutritional status score and disease activity of ulcerative colitis.

OBJECTIVE: To explore the association between the controlling nutritional status (CONUT) score and disease activity in patients with ulcerative colitis (UC). METHODS: This retrospective study enrolled patients with UC. Demographic, clinical and laboratory data were collected and compared. The CONUT score was obtained for each patient. The association between the CONUT score and laboratory parameters was analysed and the ability of the score to assess disease activity was evaluated. RESULTS: A total of 182 patients with UC were enrolled. Patients with active disease showed significantly increased inflammatory biomarkers and decreased nutritional biomarkers compared with patients in remission. Malnourished individuals had significantly elevated inflammatory biomarkers and significantly reduced haemoglobin, prealbumin and retinol-binding protein. The CONUT score was inversely correlated with haemoglobin, prealbumin, retinol-binding protein and was positively correlated with faecal calprotectin, C-reactive protein, erythrocyte sedimentation rate, neutrophil/lymphocyte ratio and platelet/lymphocyte ratio. The area under the receiver operating characteristic curve was 0.655 (95% confidence interval, 0.557-0.752). The optimal cut-off value was 1.5 points, with a sensitivity of 75.7% and a specificity of 50.0%. CONCLUSION: The CONUT score may evaluate the inflammatory response and nutritional status of UC patients, so it could be a potential biomarker to assess disease activity in UC.

Retinal Responses to Visual Stimuli in Interphotoreceptor Retinoid Binding-Protein Knock-Out Mice.

Interphotoreceptor retinoid-binding protein (IRBP) is an abundant glycoprotein in the subretinal space bound by the photoreceptor (PR) outer segments and the processes of the retinal pigmented epithelium (RPE). IRBP binds retinoids, including 11-cis-retinal and all-trans-retinol. In this study, visual function for demanding visual tasks was assessed in IRBP knock-out (KO) mice. Surprisingly, IRBP KO mice showed no differences in scotopic critical flicker frequency (CFF) compared to wildtype (WT). However, they did have lower photopic CFF than WT. IRBP KO mice had reduced scotopic and photopic acuity and contrast sensitivity compared to WT. IRBP KO mice had a significant reduction in outer nuclear layer (ONL) thickness, PR outer and inner segment, and full retinal thickness (FRT) compared to WT. There were fewer cones in IRBP KO mice. Overall, these results confirm substantial loss of rods and significant loss of cones within 30 days. Absence of IRBP resulted in cone circuit damage, reducing photopic flicker, contrast sensitivity, and spatial frequency sensitivity. The c-wave was reduced and accelerated in response to bright steps of light. This result also suggests altered retinal pigment epithelium activity. There appears to be a compensatory mechanism such as higher synaptic gain between PRs and bipolar cells since the loss of the b-wave did not linearly follow the loss of rods, or the a-wave. Scotopic CFF is normal despite thinning of ONL and reduced scotopic electroretinogram (ERG) in IRBP KO mice, suggesting either a redundancy or plasticity in circuits detecting (encoding) scotopic flicker at threshold even with substantial rod loss.

Effect of three enteral diets with different protein contents on protein metabolism in critically ill infants: a randomized controlled trial.

INTRODUCTION: The optimal intake to improve protein metabolism without producing adverse effects in seriously ill infants has yet to be established. The aim of our study was to analyse whether an increased protein intake delivered through enteral nutrition would be associated with an improvement in nitrogen balance and serum protein levels in critically ill infants. METHODS: We conducted a multicentre, prospective randomized controlled trial (December 2016-June 2019). The sample consisted of critically ill infants receiving enteral nutrition assigned randomly to 3 protein content groups: standard diet (1.7 g/dL), protein-enriched diet (2.7 g/dL) and high protein-enriched diet (5.1 g/dL). Blood and urine tests were performed, and we assessed nitrogen balance at baseline and at 3-5 days of the diet. We analysed variations in nitrogen balance and serum protein levels (total protein, albumin, transferrin, prealbumin, and retinol-binding protein) throughout the study period. RESULTS: Ninety-nine infants (33 per group) completed the study. We did not find any differences were between groups in demographic characteristics, severity scores or prescribed medications, except for corticosteroids, administered in a higher proportion of patients in the third group. We observed significant increases in prealbumin and retinol-binding protein levels in patients receiving the protein-enriched and high protein-enriched diets at 3-5 days compared to baseline. The nitrogen balance increased in all groups, but the differences were not significant in the high protein-enriched group. There were no differences in gastrointestinal tolerance. Patients fed high protein-enriched formula had higher levels of serum urea, with a higher incidence of hyperuraemia in this group. CONCLUSION: Enteral administration of higher amounts of protein improves serum protein levels in critically ill children. A protein intake of 2.2 g/kg/day is generally safe and well tolerated, whereas an intake of 3.4 g/kg/day may produce hyperuraemia in some patients.

A high fat diet fosters elevated bisretinoids.

High dietary fat intake is associated with metabolic dysregulation, but little is known regarding the effects of a high fat diet (HFD) on photoreceptor cell functioning. We explored the intersection of an HFD and the visual cycle adducts that form in photoreceptor cells by nonenzymatic reactions. In black C57BL/6J mice and albino C57BL/6Jc2j mice raised on an HFD until age 3, 6, or 12 months, chromatographically quantified bisretinoids were increased relative to mice on a standard diet. In vivo measurement of fundus autofluorescence, the source of which is bisretinoid, also revealed a significant increase in the HFD mice. Additionally, mice provided with a diet high in fat presented with elevated retinol-binding protein 4, the protein responsible for transporting retinol in plasma. Vitamin A was elevated in plasma although not in ocular tissue. Bisretinoids form in photoreceptor cell outer segments by random reactions of retinaldehyde with phosphatidylethanolamine. We found that the latter phospholipid was significantly increased in mice fed an HFD versus mice on a control diet. In leptin-deficient ob/ob mice, a genetic model of obesity, plasma levels of retinol-binding protein 4 were higher but bisretinoids in retina were not elevated. Photoreceptor cell viability measured as outer nuclear layer thickness was reduced in the ob/ob mice relative to WT. The accelerated formation of bisretinoid we observed in diet-induced obese mice is related to the high fat intake and to increased delivery of vitamin A to the visual cycle.

Retinol-binding protein-hijacking nanopolyplex delivering siRNA to cytoplasm of hepatic stellate cell for liver fibrosis alleviation.

Activated hepatic stellate cell (aHSC) is mainly responsible for deposition of extracellular collagen matrix that causes liver fibrosis. Although several siRNAs adequately inhibited HSC activation in vitro, they were demonstrated poor RNAi efficiency in vivo. Developing HSC-targeting and cytoplasmic delivery nanocarrier is highly essential to acquire a desirable siRNA therapeutic index for anti-liver fibrosis. Here, we developed a unique crosslinking nanopolyplex (called T-C-siRNA) modified by vitamin A (VA) with the well-designed natures, including the negative charge, retinol-binding protein (RBP) hijacking, and cytoplasmic siRNA release in response to ROS and cis diol molecules. The nanopolyplex was given a yolk-shell-like shape, camouflage ability in blood, and HSC-targeting capability by hijacking the endogenous ligand RBP via surface VA. PDGFR-ß siRNA (siPDGFR-ß) supplied via T-C-siPDGFR-ß nanopolyplex dramatically reduced HSC activation and its production of pro-fibrogenic proteins in vitro and in vivo. Furthermore, T-C-siPDGFR-ß nanopolyplex effectively alleviated CCl4-induced liver injury, decreased hepatic collagen sediment, and recovered liver function in mice. This study provides a sophisticated method for HSC-targeting cytoplasmic RNA delivery using endogenous ligand hijacking and dual sensitivity of ROS and cis diol compounds.

Comparative molecular dynamics study on the features of binding and non-binding modes of retinoic acid in cellular retinol-binding protein (I).

Retinoids play crucial roles in various biological processes by interacting with their carrier proteins such as cellular retinol-binding protein (CRBP). Understanding the molecular interactions between retinoids and CRBP enables their pharmacological and biomedical applications. Experimentally, CRBP(I) does not bind to retinoic acid, but when arginine is introduced into 108th residue instead of glutamine (Q108R), it binds to retinoic acid. Here, molecular dynamics simulations were performed to understand the differences in the microscopic and dynamic behaviors of the non-binding wild-type CRBP(I)-retinoic acid and binding Q108R variant-retinoic acid complexes. The ligand RMSD and RMSF, the binding poses of binding motif amino acids, and the number of hydrogen bonds and salt-bridges revealed the relative instability of the non-binding complex. In particular, the ligand's terminal group showed very different dynamics and interactions. So far, most studies have focused on the binding characteristics of retinoids, but the features of their non-binding modes have not been studied well. This study provides some structural insights into the non-binding modes of a retinoid in CRBP, which may be applicable in retinoid-based drug discovery and protein engineering through computational modeling.

Retinol binding protein 4 promotes the phenotypic transformation of vascular smooth muscle cells under high glucose condition via modulating RhoA/ROCK1 pathway.

Phenotypic switch of vascular smooth muscle cells (VSMCs) contributes to the pathogenesis of atherosclerosis (AS). High level of retinol binding protein 4 (RBP4) is regarded as a risk factor in cardiac-cerebral vascular disease. This study is performed to clarify the biological function of RBP4 in modulating the phenotypic switch of VSMCs induced via RhoA/ROCK1 signaling pathway. In vivo experiment, all the rats were divided into control group (NC), diabetic group (DM) and diabetic atherosclerosis group (DAS). The expressions of biochemical indicators, RhoA and Rho associated coiled-coil containing protein kinase 1 (ROCK1) were detected. In vitro experiment, VSMCs were cultured under high glucose condition, and ectogenic RBP4, HA-1100, rapamycin, or 3-methyladenine (3-MA) were supplemented to treat the VSMCs, respectively. The proliferation and migration of VSMCs were evaluated. The regulatory relationship between RBP4 and ROCK1 was predicted by bioinformatics analysis, and validated by qRT-PCR and Western blot. The regulatory effects of RBP4 on contractile phenotypic markers such as calponin, MYH11, α-SMA and autophagy markers including LC3II, LC3I, and Beclin-1 as well as mTOR were also detected. Moreover, VSMCs were cultured exposed to ROCK1 overexpressed plasmid or short hairpin RNA (shRNA), the proliferation and migration of VSMCs were evaluated and the regulatory effects of RhoA/ROCK1 signaling pathway on contractile phenotypic markers and autophagy markers were also detected. In vivo, RhoA, ROCK1, and mTOR were highly expressed in the rats intraperitoneally injected with RBP4. In vitro, the expressions of calponin, MYH11, α-SMA, LC3II, LC3I, and Beclin-1 were decreased in VSMCs treated with ROCK1-OA under high glucose condition, conversely, the expressions were increased in VSMCs exposed to ROCK1-shRNA. After incubated with rapamycin additionally, the expressions of calponin, MYH11, α-SMA, LC3II/I and Beclin-1 were up-regulated and the expression of p-mTOR was decreaed in VSMCs of HG+ROCK1-OA. Conversely, after incubated with 3-MA additionally, the expressions of calponin, MYH11, α-SMA, LC3II/I and Beclin-1 were down-regulated and the expression of p-mTOR was elevated in VSMCs of HG+ROCK1-shRNA. Ectogenic RBP4 facilitated high glucose-induced proliferation and migration of VSMCs, and it repressed the expression of calponin, MYH11, α-SMA, LC3II/I, and Beclin-1 in VSMCs. As expected, ROCK1 inhibit or counteracted the biological effects of RBP4 on VSMCs. In addition, the expressions of contractile phenotypic markers, LC3II/I, and Beclin-1 were promoted and mTOR were decreased after the VSMCs treated with autophagy agonist, whereas no significant difference was observed in the expressions of ROCK1, RhoA. RBP4 is an injurious factor in the pathogenesis of diabetic AS, and it promotes the phenotypic switch of VSMCs via activating RhoA/ROCK1 pathway and inhibiting autophagy.

An ancient bacterial gene set the stage for human sight.

Some 500 million years ago, early vertebrates acquired bacterial DNA that gave rise to a key vision gene.