Association of KIR gene polymorphisms with COVID-19 disease.

BACKGROUND: Natural killer (NK) cells play an essential role against viruses. NK cells express killer cell immunoglobulin-like receptors (KIRs) which regulate their activity and function. The polymorphisms in KIR haplotypes confer differential viral susceptibility and disease severity caused by infections. We investigated the association between KIR genes and COVID-19 disease severity. METHODS: 424 COVID-19 positive patients were divided according to their disease severity into mild, moderate and severe. KIR genes were genotyped using next generation sequencing (NGS). Association between KIR genes and COVID-19 disease severity was conducted and significant correlations were reported. RESULTS: In the COVID-19 patients, KIR Bx genotype was more common than AA genotype. The Bx genotype was found more frequently in patients with mild disease, while in severe disease the AA genotype was more common than the Bx genotype. The KIR2DS4 gene carried the highest risk for severe COVID-19 infection (OR 8.48, pc= 0.0084) followed by KIR3DL1 (OR 7.61, pc= 0.0192). CONCLUSIONS: Our findings suggest that KIR2DS4 and KIR3DL1 genes carry risk for severe COVID-19 disease.

Interferon-induced transmembrane protein-3 genetic variant rs12252 is associated with COVID-19 mortality.

Interferon-induced membrane proteins (IFITM) 3 gene variants are known risk factor for severe viral diseases. We examined whether IFITM3 variant may underlie the heterogeneous clinical outcomes of SARS-CoV-2 infection-induced COVID-19 in large Arab population. We genotyped 880 Saudi patients; 93.8% were PCR-confirmed SARS-CoV-2 infection, encompassing most COVID-19 phenotypes. Mortality at 90 days was 9.1%. IFITM3-SNP, rs12252-G allele was associated with hospital admission (OR = 1.65 [95% CI; 1.01-2.70], P = 0.04]) and mortality (OR = 2.2 [95% CI; 1.16-4.20], P = 0.01). Patients less than 60 years old had a lower survival probability if they harbor this allele (log-rank test P = 0.002). Plasma levels of IFNγ were significantly lower in a subset of patients with AG/GG genotypes than patients with AA genotype (P = 0.00016). Early identification of these individuals at higher risk of death may inform precision public health response.

Role of sphingolipid metabolism in neurodegeneration.

Sphingolipids are a class of lipids highly enriched in the central nervous system (CNS), which shows great diversity and complexity, and has been implicated in CNS development and function. Alterations in sphingolipid metabolism have been described in multiple diseases, including those affecting the central nervous system (CNS). In this review, we discuss the role of sphingolipid metabolism in neurodegeneration, evaluating its direct roles in neuron development and health, and also in the induction of neurotoxic activities in CNS-resident astrocytes and microglia in the context of neurologic diseases such as multiple sclerosis and Alzheimer's disease. Finally, we focus on the metabolism of gangliosides and sphingosine-1-phosphate, its contribution to the pathogenesis of neurologic diseases, and its potential as a candidate target for the therapeutic modulation of neurodegeneration.

PDCD6IP, encoding a regulator of the ESCRT complex, is mutated in microcephaly.

Primary microcephaly (PM) is a highly heterogeneous neurodevelopmental disorder with many contributing risk genes and loci identified to date. We report a consanguineous family with PM, intellectual disability and short stature. Using whole exome sequencing, we identified a homozygous frameshift variant in programmed cell death 6 interacting protein (PDCD6IP, c.154_158dup; p.Val54Profs*18). This gene, PDCD6IP, plays an important role in the endosomal sorting complexes required for transport (ESCRT) pathway in the abscission stage of cytokinesis and apoptosis, and is required for normal brain development in mice. The clinical features observed in our patient were similar to the phenotypes observed in mouse and zebrafish models of PDCD6IP mutations in previous studies. This study provides evidence that clinical manifestations of PDCD6IP mutations as seen in our patients with PM and ID may be a novel cause for neurodevelopmental disorders.

ADAMTS19-associated heart valve defects: Novel genetic variants consolidating a recognizable cardiac phenotype.

Recently, ADAMTS19 was identified as a novel causative gene for autosomal recessive heart valve disease (HVD), affecting mainly the aortic and pulmonary valves. Exome sequencing and data repository (CentoMD) analyses were performed to identify patients with ADAMTS19 variants (two families). A third family was recognized based on cardiac phenotypic similarities and SNP array homozygosity. Three novel loss of function (LoF) variants were identified in six patients from three families. Clinically, all patients presented anomalies of the aortic/pulmonary valves, which included thickening of valve leaflets, stenosis and insufficiency. Three patients had (recurrent) subaortic membrane, suggesting that ADAMTS19 is the first gene identified related to discrete subaortic stenosis. One case presented a bi-commissural pulmonary valve. All patients displayed some degree of atrioventricular valve insufficiency. Other cardiac anomalies included atrial/ventricular septal defects, persistent ductus arteriosus, and mild dilated ascending aorta. Our findings confirm that biallelic LoF variants in ADAMTS19 are causative of a specific and recognizable cardiac phenotype. We recommend considering ADAMTS19 genetic testing in all patients with multiple semilunar valve abnormalities, particularly in the presence of subaortic membrane. ADAMTS19 screening in patients with semilunar valve abnormalities is needed to estimate the frequency of the HVD related phenotype, which might be not so rare.

A missense mutation in the CRBN gene that segregates with intellectual disability and self-mutilating behaviour in a consanguineous Saudi family.

BACKGROUND: Autosomal-recessive non-syndromic intellectual disability (ARNS-ID) is an aetiologically heterogeneous disorder. Although little is known about the function of human cereblon (CRBN), its relationship to mild cognitive deficits suggests that it is involved in the basic processes of human memory and learning. OBJECTIVES: We aim to identify the genetic cause of intellectual disability and self-mutilation in a consanguineous Saudi family with five affected members. METHODS: Clinical whole-exome sequencing was performed on the proband patient, and Sanger sequencing was done to validate and confirm segregation in other family members. RESULTS: A missense variant (c. 1171T>C) in the CRBN gene was identified in five individuals with severe intellectual disability (ID) in a consanguineous Saudi family. The homozygous variant was co-segregating in the family with the phenotype of severe ID, seizures and self-mutilating behaviour. The missense mutation (p.C391R) reported here results in the replacement of a conserved cysteine residue by an arginine in the CULT (cereblon domain of unknown activity, binding cellular ligands and thalidomide) domain of CRBN, which contains a zinc-binding site. CONCLUSIONS: These findings thus contribute to a growing list of ID disorders caused by CRBN mutations, broaden the spectrum of phenotypes attributable to ARNS-ID and provide new insight into genotype-phenotype correlations between CRBN mutations and the aetiology of ARNS-ID.

Association between human leukocyte antigen alleles and COVID-19 disease severity.

BACKGROUND: the human leukocyte antigen (HLA) loci have been widely characterized to be associated with viral infectious diseases. Several studies including various ethnic groups and populations suggested associations between certain HLA alleles and SARS-CoV-2 infection. Despite the numerous associations identified, the role of HLA polymorphisms in determining the individual response to SARS-CoV-2 infection is controversial among different Saudi populations. METHOD: Here, we performed HLA typing by next-generation sequencing to investigate if variations in polymorphic HLA genes are linked to COVID-19 severity in the Saudi population. Namely, we analyzed HLA loci at allele level in 575 Saudi patients with SARS-CoV-2 infection. HLA class I and class II frequencies in patients were compared with allele frequency data from healthy Saudi population. RESULTS: in our cohort HLA-A* 02:01:01 G was associated with mild disease but was not associated with moderate and severe disease. HLA-B* 51:01:01 G was protective from severe disease while HLA-B* 50:01:01 G, HLA-C* 06:02:01 G and HLA-DRB1 * 07:01:01 G were associated with risk to severe disease as well as the total COVID-19 cohort. HLA-DRB1 * 15:01:01 G was associated with risk to all severity groups. CONCLUSION: in conclusion, we found significant associations between HLA alleles and COVID-19 disease severity in Saudis. Further studies are warranted to include HLA typing in the workup for any new COVID-19 patients.

Case report: A founder UGDH variant associated with developmental epileptic encephalopathy in Saudi Arabia.

Congenital disorders of glycosylation (CDG) are a group of more than 100 rare genetic disorders characterized by impaired glycosylation of proteins and lipids. The clinical presentation of CDG varies tremendously, from single-organ to multi-organ involvement and from prenatal death to a normal adult phenotype. In this case study, we report a large consanguineous family with multiple children suffering from cerebral palsy, seizure, developmental and epileptic encephalopathy, and global developmental delay. Whole-exome sequencing (WES) analysis revealed a homozygous variant in the UDP-glucose dehydrogenase (UGDH) gene (c.950G>A; p.R317Q) which segregates with the familial phenotype with a plausible autosomal recessive mode of inheritance, indicating a potential disease-causing association. The UGDH gene encodes the UDP-glucose dehydrogenase, a key enzyme in the synthesis of specific extracellular matrix constituents (proteoglycans and glycolipids) involved in neural migration and connectivity during early brain development. Many pathogenic mutations of UGDH have been reported in recent literature works. However, the variant identified in this study has been observed only in the Saudi population (13 families) and not in any other ethnic background, suggesting that it may be an ancient founder mutation.

Tween 80-Based Self-Assembled Mixed Micelles Boost Valsartan Transdermal Delivery.

Valsartan (Val) is an important antihypertensive medication with poor absorption and low oral bioavailability. These constraints are due to its poor solubility and dissolution rate. The purpose of this study was to optimize a mixed micelle system for the transdermal delivery of Val in order to improve its therapeutic performance by providing prolonged uniform drug levels while minimizing drug side effects. Thin-film hydration and micro-phase separation were used to produce Val-loaded mixed micelle systems. A variety of factors, including the surfactant type and drug-to-surfactant ratio, were optimized to produce micelles with a low size and high Val entrapment efficiency (EE). The size, polydispersity index (PDI), zeta potential, and drug EE of the prepared micelles were all measured. The in vitro drug release profiles were assessed using dialysis bags, and the permeation through abdominal rat skin was assessed using a Franz diffusion cell. All formulations had high EE levels exceeding 90% and low particle charges. The micellar sizes ranged from 107.6 to 191.7 nm, with average PDI values of 0.3. The in vitro release demonstrated a uniform slow rate that lasted one week with varying extents. F7 demonstrated a significant (p < 0.01) transdermal efflux of 68.84 ± 3.96 µg/cm2/h through rat skin when compared to the control. As a result, the enhancement factor was 16.57. In summary, Val-loaded mixed micelles were successfully prepared using two simple methods with high reproducibility, and extensive transdermal delivery was demonstrated in the absence of any aggressive skin-modifying enhancers.

TGFBR1 Variants Can Associate with Non-Syndromic Congenital Heart Disease without Aortopathy.

BACKGROUND: Congenital heart diseases (CHD) are the most common congenital malformations in newborns and remain the leading cause of mortality among infants under one year old. Molecular diagnosis is crucial to evaluate the recurrence risk and to address future prenatal diagnosis. Here, we describe two families with various forms of inherited non-syndromic CHD and the genetic work-up and resultant findings. METHODS: Next-generation sequencing (NGS) was employed in both families to uncover the genetic cause. In addition, we performed functional analysis to investigate the consequences of the identified variants in vitro. RESULTS: NGS identified possible causative variants in both families in the protein kinase domain of the TGFBR1 gene. These variants occurred on the same amino acid, but resulted in differently substituted amino acids (p.R398C/p.R398H). Both variants co-segregate with the disease, are extremely rare or unique, and occur in an evolutionary highly conserved domain of the protein. Furthermore, both variants demonstrated a significantly altered TGFBR1-smad signaling activity. Clinical investigation revealed that none of the carriers had (signs of) aortopathy. CONCLUSION: In conclusion, we describe two families, with various forms of inherited non-syndromic CHD without aortopathies, associated with unique/rare variants in TGFBR1 that display altered TGF-beta signaling. These findings highlight involvement of TGFBR1 in CHD, and warrant consideration of potential causative TGFBR1 variants also in CHD patients without aortopathies.

Consanguinity and Congenital Heart Disease Susceptibility: Insights into Rare Genetic Variations in Saudi Arabia.

Congenital heart disease (CHD) encompasses a wide range of structural defects of the heart and, in many cases, the factors that predispose an individual to disease are not well understood, highlighting the remarkable complexity of CHD etiology. Evidence of familial aggregation of CHD has been demonstrated in different communities and for different cardiac lesions. Consanguinity, particularly among first cousins, is an added risk factor for these families, particularly in societies where it is considered a common cultural practice, as confirmed in previous studies conducted in Saudi Arabia and other countries. Through comprehensive genetic testing of affected families, we have been able to better understand the genetic basis of the various cardiac lesions and to delineate the molecular mechanisms involved in cardiac morphogenesis. In this review, we discuss the epidemiology and genetics of CHD in consanguineous populations focusing on Saudi Arabia as an extensive study model to address current advances and challenges in the clinical genetic diagnosis and prevention of CHD.

Correlation between ABO Blood Group Phenotype and the Risk of COVID-19 Infection and Severity of Disease in a Saudi Arabian Cohort.

BACKGROUND: Disease severity among patients infected with SARS-CoV-2 varies remarkably. Preliminary studies reported that the ABO blood group system confers differential viral susceptibility and disease severity caused by SARS-CoV-2. Thus, differences in ABO blood group phenotypes may partly explain the observed heterogeneity in COVID-19 severity patterns, and could help identify individuals at increased risk. Herein, we explored the association between ABO blood group phenotypes and COVID-19 susceptibility and severity in a Saudi Arabian cohort. METHODS: In this retrospective cohort study, we performed ABO typing on a total of 373 Saudi patients infected with SARS-CoV-2 and conducted association analysis between ABO blood group phenotype and COVID-19 infection severity. We then performed gender-stratified analysis by dividing the participating patients into two groups by gender, and classified them according to age. RESULTS: The frequencies of blood group phenotypes A, B, AB and O were 27.3, 23.6, 5.4 and 43.7%, respectively. We found that blood group phenotype O was associated with a lower risk of testing positive for COVID-19 infection (OR 0.76 95% CI 0.62-0.95, p = 0.0113), while blood group phenotype B was associated with higher odds of testing positive (OR 1.51 95% CI 1.17-1.93, p = 0.0009). However, blood group phenotype B was associated with increased risk in the mild and moderate group but not the severe COVID-19 infection group. Blood group phenotype O was protective in all severity groups. CONCLUSION: Our findings provide evidence that blood group phenotype B is a risk for COVID-19 disease while blood group phenotype O is protective from COVID-19 infection. However, further studies are necessary to validate these associations in a larger sample size and among individuals of different ethnic groups.

The Role of the Disrupted Podosome Adaptor Protein (SH3PXD2B) in Frank-Ter Haar Syndrome.

Frank-Ter Haar syndrome (FTHS), sometimes referred to as Ter Haar syndrome, is a rare hereditary disorder that manifests in skeletal, cardiac, and ocular anomalies, including hypertelorism, glaucoma, prominent eyes, and facial abnormalities. In this study, we performed whole-exome sequencing (WES) to identify the genetic component responsible for the phenotype of the index patient, a male infant born to a consanguineous family from Saudi Arabia. The analysis revealed a homozygous missense variant, c.280C>G, in the SH3PXD2B gene, which cosegregates with the familial phenotype with a plausible autosomal-recessive mode of inheritance, indicating a potential disease-causing association. The SH3PXD2B gene encodes a TKS4 podosome adaptor protein that regulates the epidermal growth factor signaling pathway. This study validates the critical function of the TKS4 podosome protein by suggesting a common mechanism underlying the pathogenesis of FTHS.

Analysis of chronic kidney disease patients by targeted next-generation sequencing identifies novel variants in kidney-related genes.

Despite the enormous economic and societal burden of chronic kidney disease (CKD), its pathogenesis remains elusive, impeding specific diagnosis and targeted therapy. Herein, we sought to elucidate the genetic causes of end-stage renal disease (ESRD) and identify genetic variants associated with CKD and related traits in Saudi kidney disease patients. We applied a genetic testing approach using a targeted next-generation sequencing gene panel including 102 genes causative or associated with CKD. A total of 1,098 Saudi participants were recruited for the study, including 534 patients with ESRD and 564 healthy controls. The pre-validated NGS panel was utilized to screen for genetic variants, and then, statistical analysis was conducted to test for associations. The NGS panel revealed 7,225 variants in 102 sequenced genes. Cases had a significantly higher number of confirmed pathogenic variants as classified by the ClinVar database than controls (i.e., individuals with at least one allele of a confirmed pathogenic variant that is associated with CKD; 279 (0.52) vs. 258 (0.45); p-value = 0.03). A total of 13 genetic variants were found to be significantly associated with ESRD in PLCE1, CLCN5, ATP6V1B1, LAMB2, INVS, FRAS1, C5orf42, SLC12A3, COL4A6, SLC3A1, RET, WNK1, and BICC1, including four novel variants that were not previously reported in any other population. Furthermore, studies are necessary to validate these associations in a larger sample size and among individuals of different ethnic groups.

Reply to Zandi, M.; Soltani, S. Comment on "Alfassam et al. Development of a Colorimetric Tool for SARS-CoV-2 and Other Respiratory Viruses Detection Using Sialic Acid Fabricated Gold Nanoparticles. Pharmaceutics 2021, 13, 502".

When COVID-19 was first announced back in 2019, there were vast number of attempts to halt the progression of the SARS-CoV-2 virus once and for all [...].

Fast-Dissolving Nifedipine and Atorvastatin Calcium Electrospun Nanofibers as a Potential Buccal Delivery System.

Geriatric patients are more likely to suffer from multiple chronic diseases that require using several drugs, which are commonly ingested. However, to enhance geriatric patients' convenience, the electrospun nanofiber system was previously proven to be a successful alternative for the existing oral dosage forms, i.e., tablets and capsules. These nanofibers prepared either as single- or multi-layered fibers could hold at least one active compound in each layer. They might also be fabricated as ultra-disintegrated fibrous films for oral cavity administration, i.e., buccal or sublingual, to improve the bioavailability and intake of the administered drugs. Therefore, in this work, a combination of nifedipine and atorvastatin calcium, which are frequently prescribed for hypertension and hyperlipidemia patients, respectively, was prepared in a coaxial electrospinning system for buccal administration. Scanning electron microscopy image showed the successful preparation of smooth, non-beaded, and non-porous surfaces of the drug-loaded nanofibers with an average fiber diameter of 968 ± 198 nm. In contrast, transmission electron microscopy distinguished the inner and outer layers of those nanofibers. The disintegration of the drug-loaded nanofibers was ≤12 s, allowing the rapid release of nifedipine and atorvastatin calcium to 61% and 47%, respectively, after 10 min, while a complete drug release was achieved after 120 min. In vitro, a drug permeation study using Franz diffusion showed that the permeation of both drugs from the core-shell nanofibers was enhanced significantly (p < 0.05) compared to the drugs in a solution form. In conclusion, the development of drug-loaded nanofibers containing nifedipine and atorvastatin calcium can be a potential buccal delivery system.

Development of Epirubicin-Loaded Biocompatible Polymer PLA-PEG-PLA Nanoparticles: Synthesis, Characterization, Stability, and In Vitro Anticancerous Assessment.

Epirubicin (EPI) is an anti-cancerous chemotherapeutic drug that is an effective epimer of doxorubicin with less cardiotoxicity. Although EPI has fewer side effects than its analog, doxorubicin, this study aims to develop EPI nanoparticles as an improved formula of the conventional treatment of EPI in its free form. METHODS: In this study, EPI-loaded polymeric nanoparticles (EPI-NPs) were prepared by the double emulsion method using a biocompatible poly (lactide) poly (ethylene glycol) poly(lactide) (PLA-PEG-PLA) polymer. The physicochemical properties of the EPI-NPs were determined by dynamic light scattering (DLS), transmission electron microscopy (TEM), differential scanning calorimetry (DSC), entrapment efficiency and stability studies. The effect of EPI-NPs on cancer cells was determined by high throughput imaging and flow cytometry. RESULTS: The synthesis process resulted in monodisperse EPI-NPs with a size of 166.93 ± 1.40 nm and an elevated encapsulation efficiency (EE) of 88.3%. In addition, TEM images revealed the spherical uniformness of EPI-NPs with no aggregation, while the cellular studies presented the effect of EPI-NPs on MCF-7 cells' viability; after 96 h of treatment, the MCF-7 cells presented considerable apoptotic activity. The stability study showed that the EPI-NPs remained stable at room temperature at physiological pH for over 30 days. CONCLUSION: EPI-NPs were successfully encapsulated within a highly stable biocompatible polymer with minimal loss of the drug. The used polymer has low cytotoxicity and EPI-NPs induced apoptosis in estrogen-positive cell line, making them a promising, safe treatment for cancer with less adverse side effects.

Novel Autosomal Recessive Splice-Altering Variant in PRKD1 Is Associated with Congenital Heart Disease.

Congenital heart defects (CHDs) are the most common types of birth defects, and global incidence of CHDs is on the rise. Despite the prevalence of CHDs, the genetic determinants of the defects are still in the process of being identified. Herein, we report a consanguineous Saudi family with three CHD affected daughters. We used whole exome sequencing (WES) to investigate the genetic cause of CHDs in the affected daughters. We found that all affected individuals were homozygous for a novel splice-altering variant (NM_001330069.1: c.265-1G>T) of PRKD1, which encodes a calcium/calmodulin-dependent protein kinase in the heart. The homozygous variant was found in the affected patients with Pulmonary Stenosis (PS), Truncus Arteriosis (TA), and Atrial Septal Defect (ASD). Based on the family's pedigree, the variant acts in an autosomal recessive manner, which makes it the second autosomal recessive variant of PRKD1 to be identified with a link to CHDs, while all other previously described variants act dominantly. Interestingly, the father of the affected daughters was also homozygous for the variant, though he was asymptomatic of CHDs himself. Since both of his sisters had CHDs as well, this raises the possibility that the novel PRKD1 variant may undergo autosomal recessive inheritance mode with gender limitation. This finding confirms that CHD can be associated with both dominant and recessive mutations of the PRKD1 gene, and it provides a new insight to genotype-phenotype association between PRKD1 and CHDs. To our knowledge, this is the first report of this specific PRKD1 mutation associated with CHDs.

Development of a Colorimetric Tool for SARS-CoV-2 and Other Respiratory Viruses Detection Using Sialic Acid Fabricated Gold Nanoparticles.

Sialic acid that presents on the surface of lung epithelial cells is considered as one of the main binding targets for many respiratory viruses, including influenza and the current coronavirus (SARS-CoV-2) through the viral surface protein hemagglutinin. Gold nanoparticles (Au NPs) are extensively used in the diagnostic field owing to a phenomenon known as 'surface plasmonic resonance' in which the scattered light is absorbed by these NPs and can be detected via UV-Vis spectrophotometry. Consequently, sialic acid conjugated Au NPs (SA-Au NPs) were utilized for their plasmonic effect against SARS-CoV-2, influenza B virus, and Middle-East respiratory syndrome-related coronavirus (MERS) in patients' swab samples. The SA-Au NPs system was prepared by a one-pot synthesis method, through which the NPs solution color changed from pale yellow to dark red wine color, indicting its successful preparation. In addition, the SA-Au NPs had an average particle size of 30 ± 1 nm, negative zeta potential (-30 ± 0.3 mV), and a UV absorbance of 525 nm. These NPs have proven their ability to change the color of the NPs solutions and patients' swabs that contain SARS-CoV-2, influenza B, and MERS viruses, suggesting a rapid and straightforward detection tool that would reduce the spread of these viral infections and accelerate the therapeutic intervention.

A cell-based drug delivery platform for treating central nervous system inflammation.

Mesenchymal stem cells (MSCs) are promising candidates for the development of cell-based drug delivery systems for autoimmune inflammatory diseases, such as multiple sclerosis (MS). Here, we investigated the effect of Ro-31-8425, an ATP-competitive kinase inhibitor, on the therapeutic properties of MSCs. Upon a simple pretreatment procedure, MSCs spontaneously took up and then gradually released significant amounts of Ro-31-8425. Ro-31-8425 (free or released by MSCs) suppressed the proliferation of CD4+ T cells in vitro following polyclonal and antigen-specific stimulation. Systemic administration of Ro-31-8425-loaded MSCs ameliorated the clinical course of experimental autoimmune encephalomyelitis (EAE), a murine model of MS, displaying a stronger suppressive effect on EAE than control MSCs or free Ro-31-8425. Ro-31-8425-MSC administration resulted in sustained levels of Ro-31-8425 in the serum of EAE mice, modulating immune cell trafficking and the autoimmune response during EAE. Collectively, these results identify MSC-based drug delivery as a potential therapeutic strategy for the treatment of autoimmune diseases. KEY MESSAGES: MSCs can spontaneously take up the ATP-competitive kinase inhibitor Ro-31-8425. Ro-31-8425-loaded MSCs gradually release Ro-31-8425 and exhibit sustained suppression of T cells. Ro-31-8425-loaded MSCs have more sustained serum levels of Ro-31-8425 than free Ro-31-8425. Ro-31-8425-loaded MSCs are more effective than MSCs and free Ro-31-8425 for EAE therapy.