hsa-miR-CHA2, a novel microRNA, exhibits anticancer effects by suppressing cyclin E1 in human non-small cell lung cancer cells.

Despite considerable therapeutic advancements, the global survival rate for lung cancer patients remains poor, posing challenges in developing an effective treatment strategy. In many cases, microRNAs (miRNAs) exhibit abnormal expression levels in cancers, including lung cancer. Dysregulated miRNAs often play a crucial role in the development and progression of cancer. Therefore, understanding the mechanisms underlying aberrant miRNA expression during carcinogenesis may provide crucial clues to develop novel therapeutics. In this study, we identified and cloned a novel miRNA, hsa-miR-CHA2, which is abnormally downregulated in non-small cell lung cancer (NSCLC)-derived cell lines and tissues of patients with NSCLC. Furthermore, we found that hsa-miR-CHA2 regulates the post-transcriptional levels of Cyclin E1 (CCNE1) by binding to the 3'-UTR of CCNE1 mRNA. CCNE1, a cell cycle regulator involved in the G1/S transition, is often amplified in various cancers. Notably, hsa-miR-CHA2 overexpression led to the alteration of the Rb-E2F pathway, a significant signaling pathway in the cell cycle, by targeting CCNE1 in A549 and SK-LU-1 cells. Subsequently, we confirmed that hsa-miR-CHA2 induced G1-phase arrest and exhibited an anti-proliferative effect by targeting CCNE1. Moreover, in subcutaneous xenograft mouse models, intra-tumoral injection of polyplexed hsa-miR-CHA2 mimic suppressed tumor growth and development. In conclusion, hsa-miR-CHA2 exhibited an anticancer effect by targeting CCNE1 both in vitro and in vivo. These findings suggest the potential role of hsa-miR-CHA2 as an important regulator of cell proliferation in molecular-targeted therapy for NSCLC.

Global carrier frequency and predicted genetic prevalence of patients with pathogenic sequence variants in autosomal recessive genetic neuromuscular diseases.

Genetic neuromuscular diseases are clinically and genetically heterogeneous genetic disorders that primarily affect the peripheral nerves, muscles, and neuromuscular junctions. This study aimed to identify pathogenic variants, calculate carrier frequency, and predict the genetic prevalence of autosomal recessive neuromuscular diseases (AR-NMDs). We selected 268 AR-NMD genes and analyzed their genetic variants sourced from the gnomAD database. After identifying the pathogenic variants using an algorithm, we calculated the carrier frequency and predicted the genetic prevalence of AR-NMDs. In total, 10,887 pathogenic variants were identified, including 3848 literature verified and 7039 manually verified variants. In the global population, the carrier frequency of AR-NMDs is 32.9%, with variations across subpopulations ranging from 22.4% in the Finnish population to 36.2% in the non-Finnish European population. The predicted genetic prevalence of AR-NMDs was estimated to be 24.3 cases per 100,000 individuals worldwide, with variations across subpopulations ranging from 26.5 to 41.4 cases per 100,000 individuals in the Latino/Admixed American and the Ashkenazi Jewish populations, respectively. The AR-NMD gene with the highest carrier frequency was GAA (1.3%) and the variant with the highest allele frequency was c.-32-13 T>G in GAA with 0.0033 in the global population. Our study revealed a higher-than-expected frequency of AR-NMD carriers, constituting approximately one-third of the global population, highlighting ethnic heterogeneity in genetic susceptibility.

Transcriptome analysis of skeletal muscle in dermatomyositis, polymyositis, and dysferlinopathy, using a bioinformatics approach.

Background: Polymyositis (PM) and dermatomyositis (DM) are two distinct subgroups of idiopathic inflammatory myopathies. Dysferlinopathy, caused by a dysferlin gene mutation, usually presents in late adolescence with muscle weakness, degenerative muscle changes are often accompanied by inflammatory infiltrates, often resulting in a misdiagnosis as polymyositis. Objective: To identify differential biological pathways and hub genes related to polymyositis, dermatomyositis and dysferlinopathy using bioinformatics analysis for understanding the pathomechanisms and providing guidance for therapy development. Methods: We analyzed intramuscular ribonucleic acid (RNA) sequencing data from seven dermatomyositis, eight polymyositis, eight dysferlinopathy and five control subjects. Differentially expressed genes (DEGs) were identified by using DESeq2. Enrichment analyses were performed to understand the functions and enriched pathways of DEGs. A protein-protein interaction (PPI) network was constructed, and clarified the gene cluster using the molecular complex detection tool (MCODE) analysis to identify hub genes. Results: A total of 1,048, 179 and 3,807 DEGs were detected in DM, PM and dysferlinopathy, respectively. Enrichment analyses revealed that upregulated DEGs were involved in type 1 interferon (IFN1) signaling pathway in DM, antigen processing and presentation of peptide antigen in PM, and cellular response to stimuli in dysferlinopathy. The PPI network and MCODE cluster identified 23 genes related to type 1 interferon signaling pathway in DM, 4 genes (PDIA3, HLA-C, B2M, and TAP1) related to MHC class 1 formation and quality control in PM, and 7 genes (HSPA9, RPTOR, MTOR, LAMTOR1, LAMTOR5, ATP6V0D1, and ATP6V0B) related to cellular response to stress in dysferliniopathy. Conclusion: Overexpression of genes related to the IFN1 signaling pathway and major histocompatibility complex (MHC) class I formation was identified in DM and PM, respectively. In dysferlinopathy, overexpression of HSPA9 and the mTORC1 signaling pathway genes was detected.

Association between PAI-1 Polymorphisms and Ischemic Stroke in a South Korean Case-Control Cohort.

Stroke is the second leading cause of death in the world. Approximately 80% of strokes are ischemic in origin. Many risk factors have been linked to stroke, including an increased level of plasminogen activator inhibitor-1 (PAI-1). PAI-1 levels increase and remain elevated in blood during the acute phase of ischemic stroke, which can impair fibrinolytic activity, leading to coronary artery disease and arterial thrombotic disorders. Here, we present a case-control study of 574 stroke patients and 425 controls seen for routine health examination or treatment for nonspecific dizziness, nonorganic headache, or anxiety for positive family history of stroke at the Bundang Medical Center in South Korea. Polymorphisms in PAI-1 were identified by polymerase chain reaction/restriction fragment length polymorphism analysis using genomic DNA. Specifically, three variations (-675 4G>5G, 10692T>C, and 12068G>A) were linked to a higher overall prevalence of stroke as well as a higher prevalence of certain stroke subtypes. Haplotype analyses also revealed combinations of these variations (-844G>A, -675 4G>5G, 43G>A, 9785A>G, 10692T>C, 11053T>G, and 12068G>A) that were significantly associated with a higher prevalence of ischemic stroke. To the best of our knowledge, this is the first strong evidence that polymorphic sites in PAI-1 promoter and 3'-UTR regions are associated with higher ischemic stroke risk. Furthermore, the PAI-1 genotypes and haplotypes identified here have potential as clinical biomarkers of ischemic stroke and could improve the prognosis and future management of stroke patients.

High Mean Platelet Volume Is Associated with Cerebral White Matter Hyperintensities in Non-Stroke Individuals.

PURPOSE: The mean platelet volume (MPV) is regarded as a marker for thrombosis, atherosclerosis, and inflammation in various vascular diseases. However, it still remains unclear whether plasma MPV is associated with cerebral white matter hyperintensities (WMH) and cerebral microvascular pathology in the elderly population. MATERIALS AND METHODS: We examined whether MPV level is associated with the presence of cerebral WMH on brain magnetic resonance imaging from 870 non-stroke outpatient subjects. The subjects were divided into three groups according to the consecutive level of MPV (low T1, middle T2, and high T3 MPV tertile groups). To determine the association of MPV levels with the WMH, logistic regression and receiver operating characteristic curve analyses were conducted. RESULTS: Subjects with higher MPV level were older and more likely to have hypertension, diabetes mellitus, and low renal function. Cerebral WMH were more prevalent in subjects with higher MPV level. After adjusting for confounding factors, moderate to severe cerebral WMH were significantly associated with high MPV tertile level. This association remained significant after adjusting for other cerebral vascular pathologies. T2 [odds ratio (OR): 1.49, 95% confidence interval (CI): 1.03-2.15] and T3 MPV tertile groups (OR: 1.51, 95%CI: 1.04-2.20) had more cerebral WMH lesions compared to T1 MPV tertile group. In addition, the subjects with higher Fazekas scores showed higher MPV level (p=0.020). CONCLUSION: We found that high MPV level is independently associated with cerebral WMH. This result suggests that platelet activation plays a role in the development of cerebral WMH.

The clinical significance of peripheral blood cell ratios in patients with intracranial aneurysm.

Background and objective: Inflammation is an important factor in the development of aneurysm, and has been identified as a key characteristic predictive of rupture of intracranial aneurysm (IA). However, the role of inflammatory peripheral blood cell ratios in patients with IA has not been well delineated. Methods: A total of 1,209 patients, including 1,001 with unruptured IA and 208 with ruptured IA, were enrolled in this study. Neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), platelet-to-neutrophil ratio (PNR), and platelet-to-white-blood-cell ratio (PWR) were compared between ruptured and unruptured IA. Results: Compared with the ruptured IA group, the unruptured IA group had higher PNR {median, 65.96 [interquartile range (IQR) 48.95-85.05] vs. 37.78 (IQR, 23.17-54.05); p < 0.001} and PWR [median, 36.89 (IQR 29.38-44.56) vs. 22.39 (IQR, 16.72-29.29); p < 0.001]. In multivariate analysis, PNR and PWR were independently associated with ruptured IA (p = 0.001 and p < 0.001, respectively). Unruptured IA subgroup analyses according to the PHASES scores showed that a higher PHASES score was associated with significantly higher NLR and erythrocyte sedimentation rate (p < 0.001 and p = 0.025) and lower PNR and PWR (p < 0.001 and p = 0.007). Conclusions: We demonstrated that lower PNR and PWR levels are associated with ruptured IA and a higher PHASES score. Unlike many other inflammatory markers and bioassays, peripheral blood cell ratios are inexpensive and readily available biomarkers that may be useful for risk stratification in patients with cerebral aneurysm. However, a long-term prospective study is needed to clarify this matter.

Extracellular Vesicles Released from Neprilysin Gene-Modified Human Umbilical Cord-Derived Mesenchymal Stem Cell Enhance Therapeutic Effects in an Alzheimer's Disease Animal Model.

Alzheimer's disease (AD) animal studies have reported that mesenchymal stem cells (MSCs) have therapeutic effects; however, clinical trial results are controversial. Neprilysin (NEP) is the main cleavage enzyme of ß-amyloid (Aß), which plays a major role in the pathology and etiology of AD. We evaluated whether transplantation of MSCs with NEP gene modification enhances the therapeutic effects in an AD animal model and then investigated these pathomechanisms. We manufactured NEP gene-enhanced human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) and intravenously transplanted them in Aß 1-42-injected AD animal models. We compared the differences in behavioral tests and immunohistochemical assays between four groups: normal, Aß 1-42 injection, naïve hUC-MSCs, and NEP-enhanced hUC-MSCs. Both naïve and NEP-enhanced hUC-MSC groups showed significant improvements in memory compared to the Aß 1-42 injection group. There was no significant difference between naïve and NEP-enhanced hUC-MSC groups. There was a significant decrease in Congo red, BACE-1, GFAP, and Iba-1 and a significant increase in BDNF, NeuN, and NEP in both hUC-MSC groups compared to the Aß 1-42 injection group. Among them, BDNF, NeuN, GFAP, Iba-1, and NEP showed more significant changes in the NEP-enhanced hUC-MSC group than in the naïve group. After stem cell injection, stem cells were not found. Extracellular vesicles (EVs) were equally observed in the hippocampus in the naïve and NEP-enhanced hUC-MSC groups. However, the EVs of NEP-enhanced hUC-MSCs contained higher amounts of NEP as compared to the EVs of naïve hUC-MSCs. Thus, hUC-MSCs affect AD animal models through stem cell-released EVs. Although there was no significant difference in cognitive function between the hUC-MSC groups, NEP-enhanced hUC-MSCs had superior neurogenesis and anti-inflammation properties compared to naïve hUC-MSCs due to increased NEP in the hippocampus by enriched NEP-possessing EVs. NEP gene-modified MSCs that release an increased amount of NEP within EVs may be a promising therapeutic option in AD treatment.

High Serum Levels of Resistin is Associated With Acute Cerebral Infarction.

BACKGROUND: The inflammatory process is involved in the pathogenesis of atherosclerosis and brain tissue injury following cerebral ischemia. Human resistin is a member of small cysteine-rich secreted proteins and has been implicated in inflammatory responses. This study investigated the association of serum resistin level with acute cerebral infarction (ACI). We also investigated its association with the short-term functional outcome. METHODS: This study included 106 patients with ACI and 106 age-matched and sex-matched healthy control subjects. Serum resistin level was assessed by using enzyme-linked immunosorbent sandwich assay. The association of serum resistin levels with ACI was analyzed by logistic regression analysis. RESULTS: The serum resistin level was significantly higher in patients with ACI than the control group [median (interquartile range), 35.7 ng/mL (13.0 to 70.5) ng/mL vs. 10.5 ng/ml (15.4 to 16.6), P<0.001]. Logistic regression analysis showed that serum resistin level was associated with an ACI (odds ratio=1.055, 95% confidence interval: 1.035-1.074, P<0.001). Among stroke subtypes, the serum resistin level was higher in the patients with large artery atherosclerosis than those with other subtypes (P=0.013). High resistin levels were also significantly associated with unfavorable functional outcome at discharge (odds ratio=1.043, 95% confidence interval: 1.024-1.063, P<0.001). CONCLUSIONS: This study suggests the potential association of resistin with stroke and cerebral atherosclerosis. Increased serum resistin levels were also associated with early unfavorable neurological outcome.

Therapeutic Effect of BDNF-Overexpressing Human Neural Stem Cells (F3.BDNF) in a Contusion Model of Spinal Cord Injury in Rats.

The most common type of spinal cord injury is the contusion of the spinal cord, which causes progressive secondary tissue degeneration. In this study, we applied genetically modified human neural stem cells overexpressing BDNF (brain-derived neurotrophic factor) (F3.BDNF) to determine whether they can promote functional recovery in the spinal cord injury (SCI) model in rats. We transplanted F3.BDNF cells via intrathecal catheter delivery after a contusion of the thoracic spinal cord and found that they were migrated toward the injured spinal cord area by MR imaging. Transplanted F3.BDNF cells expressed neural lineage markers, such as NeuN, MBP, and GFAP and were functionally connected to the host neurons. The F3.BDNF-transplanted rats exhibited significantly improved locomotor functions compared with the sham group. This functional recovery was accompanied by an increased volume of spared myelination and decreased area of cystic cavity in the F3.BDNF group. We also observed that the F3.BDNF-transplanted rats showed reduced numbers of Iba1- and iNOS-positive inflammatory cells as well as GFAP-positive astrocytes. These results strongly suggest the transplantation of F3.BDNF cells can modulate inflammatory cells and glia activation and also improve the hyperalgesia following SCI.

The 3'-UTR Polymorphisms in the Thymidylate Synthase (TS) Gene Associated with the Risk of Ischemic Stroke and Silent Brain Infarction.

Thymidylate synthase (TS) is a key gene involved in the repair of DNA damage and DNA synthesis that plays an important role in vascular development and recovery. In particular, TS gene polymorphisms play a major role in the progression of vascular disease and cancer metastasis. Therefore, the aim of this study was to investigate the association of three TS polymorphisms (1100T>C [rs699517], 1170A>G [rs2790], and 1494ins/del [rs151264360]) with ischemic stroke and silent brain infarction (SBI) in Koreans. A total of 1299 participants (507 stroke patients, 383 SBI patients, and 409 controls) were enrolled in the study. Genotyping of the three TS polymorphisms was performed by polymerase chain reaction-restriction fragment length polymorphism analysis. To examine the association between TS gene polymorphisms and the diseases, we performed statistical analyses, including multivariable logistic regression and Fisher's exact tests. We found that TS 1100T>C and 1170A>G genotypes were strongly associated with ischemic stroke and SBI susceptibility. More specifically, the TS 1100T>C polymorphism was associated with the likelihood of ischemic stroke (TT vs. CC: AOR = 2.151, 95% CI = 1.275-3.628, P = 0.004) and SBI (TT vs. TC+CC: AOR = 1.443, 95 % CI = 1.009-2.063, P = 0.045). In contrast, the TS 1170A > G polymorphism exhibited lower correlation with the risk of stroke (AA vs. GG: AOR = 0.284, 95% CI = 0.151-0.537, P < 0.0001) and SBI (AA vs. GG: AOR = 0.070, 95% CI = 0.016-0.298, P = 0.0002). Furthermore, we confirmed that the TS 1100T>C polymorphism was synergistic with low folic acid levels (AOR = 6.749, P < 0.0001). Altogether, these results suggest that TS 1100T>C and 1170A > G polymorphisms are associated with the risk of ischemic stroke and SBI, and our study provides the first evidence that 3'-UTR variants in TS are potential biomarkers in ischemic stroke and SBI.

Clinical impact of estradiol/testosterone ratio in patients with acute ischemic stroke.

BACKGROUND: Sex hormones may be associated with a higher incidence of ischemic stroke or stroke-related events. In observational studies, lower testosterone concentrations are associated with infirmity, vascular disease, and adverse cardiovascular risk factors. Currently, female sexual hormones are considered neuroprotective agents. The purpose of this study was to assess the role of sex hormones and the ratio of estradiol/testosterone (E/T) in patients with acute ischemic stroke (AIS). METHODS: Between January 2011 and December 2016, 146 male patients with AIS and 152 age- and sex-matched control subjects were included in this study. Sex hormones, including estradiol, progesterone, and testosterone, were evaluated in the AIS patient and control groups. We analyzed the clinical and physiological levels of sex hormones and hormone ratios in these patients. RESULTS: The E/T ratio was significantly elevated among patients in the stroke group compared to those in the control group (P = 0.001). Categorization of data into tertiles revealed that patients with the highest E/T ratio were more likely to have AIS [odds ratio (OR) 3.084; 95% Confidence interval (CI): 1.616-5.886; P < 0.001) compared with those in the first tertile. The E/T ratio was also an independent unfavorable outcome predictor with an adjusted OR of 1.167 (95% CI: 1.053-1.294; P = 0.003). CONCLUSIONS: These findings support the hypothesis that increased estradiol and reduced testosterone levels are associated with AIS in men.

MiR-10a, 27a, 34b/c, and 300 Polymorphisms are Associated with Ischemic Stroke Susceptibility and Post-Stroke Mortality.

A recent study of the ischemic stroke described the roles played by miRNAs in the downregulation of specific cell-cycle gene expression and it is thought to require the development of biomarkers for the prognostic of ischemic stroke. Here, we hypothesized that four miRNA polymorphisms (miR-10a, miR-27a, miR-34b/c, and miR-300) may affect stroke susceptibility and mortality. Blood samples were collected from 530 patients and 403 controls. Genetic polymorphisms were detected by polymerase chain reaction (PCR)-restriction fragment length polymorphism analysis and real-time PCR. We found that the miR-300 rs12894467 TC genotype and the dominant model (AOR: 2.069, p-value: 0.017; AOR: 1.931, p-value: 0.027) were significantly associated with an increased risk for the ischemic stroke subtype. In Cox proportional hazard regression models, the miR-10a rs3809783 A>T and miR-34b/c rs4938723 T>C polymorphisms were associated with the mortality rates among ischemic stroke patients. We found that a miR-300 polymorphism was associated with increased ischemic stroke susceptibility among the Korean population. Additionally, polymorphisms in miR-10a and miR-34b/c were associated with the increased or decreased mortality of ischemic stroke patients. This study marks the first report of an association between ischemic stroke and miRNA polymorphisms (miR-10aA>T, miR-27aT>C, miR-34b/cT>C, and miR-300T>C) in the Korean population.

MPG and NPRL3 Polymorphisms are Associated with Ischemic Stroke Susceptibility and Post-Stroke Mortality.

Ischemic stroke is a complicated disease which is affected by environmental factors and genetic factors. In this field, various studies using whole-exome sequencing (WES) have focused on novel and linkage variants in diverse diseases. Thus, we have investigated the various novel variants, which focused on their linkages to each other, in ischemic stroke. Specifically, we analyzed the N-methylpurine DNA glycosylase (MPG) gene, which plays an initiating role in DNA repair, and the nitrogen permease regulator-like 3 (NPRL3) gene, which is involved in regulating the mammalian target of rapamycin pathway. We took blood samples of 519 ischemic stroke patients and 417 controls. Genetic polymorphisms were detected by polymerase chain reaction (PCR), real-time PCR, and restriction fragment length polymorphism (RFLP) analysis. We found that two NPRL3 polymorphisms (rs2541618 C>T and rs75187722 G>A), as well as the MPG rs2562162 C>T polymorphism, were significantly associated with ischemic stroke. In Cox proportional hazard regression models, the MPG rs2562162 was associated with the survival of small-vessel disease patients in ischemic stroke. Our study showed that NPRL3 and MPG polymorphisms are associated with ischemic stroke prevalence and ischemic stroke survival. Taken together, these findings suggest that NPRL3 and MPG genotypes may be useful clinical biomarkers for ischemic stroke development and prognosis.

Blood Neutrophil/Lymphocyte Ratio Is Associated With Cerebral Large-Artery Atherosclerosis but Not With Cerebral Small-Vessel Disease.

Background and Objective: The blood neutrophil/lymphocyte ratio (NLR) is a marker of peripheral inflammation, with a high NLR associated with an increased risk of cardiovascular events and poor prognosis in ischemic stroke. However, few data are available on the differential impact of the blood NLR on different silent cerebral vascular pathologies, including cerebral large-artery atherosclerosis (LAA) and cerebral small-vessel disease (CSVD), in neurologically healthy individuals. Methods: We evaluated cardiovascular risk factors, brain magnetic resonance imaging (MRI), and MR angiography of 950 individuals without any neurological diseases. The study participants were divided into three groups according to NLR tertile (low, middle, and high). The presences of extracranial (EC) and intracranial (IC) atherosclerosis were considered to indicate LAA on brain MR angiography. The presences of silent lacunar infarction (SLI) and cerebral white matter hyperintensities (WMHs) were analyzed as indices of CSVD on brain MRI. Results: In univariate analysis, the high NLR tertile group showed a high prevalence of old age, hyperlipidemia, and renal dysfunction and higher fasting glucose. Multivariable logistic regression analysis revealed that indices of LAA (EC atherosclerosis [odds ratio: 1.88; 95% confidence interval: 1.14-3.09; p = 0.01] and IC atherosclerosis [odds ratio: 1.87; 95% confidence interval: 1.15-3.06; p = 0.01]) were more prevalent in the highest NLR tertile group than in the lowest NLR tertile group after adjustment for cardiovascular risk factors. However, no significant differences were found in the prevalence of CSVD indices (SLI and WMHs) among the three NLR tertile groups. Conclusions: A high NLR is associated with the development of cerebral LAA but not CSVD.

Enhancing the Therapeutic Potential of CCL2-Overexpressing Mesenchymal Stem Cells in Acute Stroke.

Although intravenous administration of mesenchymal stem cells (MSCs) is effective for experimental stroke, low engraftment and the limited functional capacity of transplanted cells are critical hurdles for clinical applications. C-C motif chemokine ligand 2 (CCL2) is associated with neurological repair after stroke and delivery of various cells into the brain via CCL2/CCR2 (CCL2 receptor) interaction. In this study, after CCL2-overexpressing human umbilical cord-derived MSCs (hUC-MSCs) were intravenously transplanted with mannitol in rats with middle cerebral arterial occlusion, we compared the differences between four different treatment groups: mannitol + CCL2-overexpressing hUC-MSCs (CCL2-MSC), mannitol + naïve hUC-MSCs (M-MSC), mannitol only, and control. At four-weeks post-transplantation, the CCL2-MSC group showed significantly better functional recovery and smaller stroke volume relative to the other groups. Additionally, we observed upregulated levels of CCR2 in acute ischemic brain and the increase of migrated stem cells into these areas in the CCL2-MSC group relative to the M-MSC. Moreover, the CCL2-MSC group displayed increased angiogenesis and endogenous neurogenesis, decreased neuro-inflammation but with increased healing-process inflammatory cells relative to other groups. These findings indicated that CCL2-overexpressing hUC-MSCs showed better functional recovery relative to naïve hUC-MSCs according to the increased migration of these cells into brain areas of higher CCR2 expression, thereby promoting subsequent endogenous brain repair.

Intracerebral Transplants of GMP-Grade Human Umbilical Cord-Derived Mesenchymal Stromal Cells Effectively Treat Subacute-Phase Ischemic Stroke in a Rodent Model.

In subacute and chronic phases of the stroke, there are no therapeutics available at present to promote functional recovery. Human umbilical cord-derived mesenchymal stromal cells (hUC-MSCs) are one of the candidate cell types for treating subacute-phase stroke. The benefits of cell-based therapy largely depend on the migratory capacity of products administered, as well as their potential for engraftment in targeted tissues and paracrine activities. Timing and delivery modes may also influence the outcomes of stem-cell therapy. Still, the functional recuperative effects of differing hUC-MSC delivery modes, about cell replacement and cell-to-cell paracrine activity levels, have yet to be clarified in subacute phases of stroke.This study was conducted to compare the therapeutic effects of various delivery routes when administering Good Manufacturing Practice (GMP)-grade hUC-MSCs in a rodent model of subacute-phase stroke. Cell aliquots (1 × 106) were given to rats as intravenous (IV) injections or intracerebral (IC) transplants 1 week after middle cerebral artery occlusion (MCAo). Transplanted rats were examined up to 7 weeks later using various behavioral tests and immunohistochemical analyses. Most IC-transplanted cells survived for short periods (i.e., <4 weeks after receipt) and gradually disappeared, whereas IV-injected cells were undetectable in the brain at the same time points (i.e., 3 days, 4 weeks, or 7 weeks after injection). Although short-lived, IC-transplanted cells effectively improved behavioral deficits, serving to reduce infarct volumes and glial scar formation, increase subventricular counts of proliferating neuroblasts, and promote cerebrovascular ingrowth in ischemic penumbra regions. IV injection, however, failed to improve behavioral function or histologic parameters during the same 7-week time frame. These findings overall suggest that IC transplantation is preferable to IV injection for delivery of hUC-MSCs during subacute phases of stroke.

Intracerebral transplantation of HLA-homozygous human iPSC-derived neural precursors ameliorates the behavioural and pathological deficits in a rodent model of ischaemic stroke.

OBJECTIVES: Human-induced pluripotent stem cells (hiPSCs) are a promising cell source for treating ischaemic stroke. Although autologous hiPSCs provide the advantage of avoiding immune rejection, their practical limitations, such as substantial amount of time and costs to generate individual iPSC lines, have hampered their widespread application in clinical settings. In this study, we investigated the therapeutic potential of neural precursor cells derived from human HLA-homozygous induced pluripotent stem cells (hiPSC-NPCs) following intracerebral transplantation into a rodent model of middle cerebral artery occlusion (MCAo). MATERIALS AND METHODS: We differentiated a GMP-grade HLA-homozygous hiPSC line (CMC-hiPSC-004) into neural precursor cells for transplantation into rats at the subacute stage of ischaemic stroke (ie at 7 days after the induction of MCAo). To investigate functional recovery, the transplanted animals were subjected to five behavioural tests, namely the rotarod, stepping, mNSS, staircase and apomorphine-induced rotation tests, for up to 12 weeks, followed by histological analyses. RESULTS: We observed that the hiPSC-NPC transplantation produced significant behavioural improvements. At 12 weeks post-transplantation, a high proportion of transplanted cells survived and had differentiated into MAP2+ mature neurons, GABAergic neurons and DARPP32+ medium spiny neurons. The transplanted cells formed neuronal connections with striatal neurons in the host brain. In addition, hiPSC-NPC transplantation gave rise to enhanced endogenous repair processes, including decreases of post-stroke neuroinflammation and glial scar formation and an increase of proliferating endogenous neural stem cells in the subventricular zone as well as the perilesional capillary networks. CONCLUSIONS: These results strongly suggest that HLA-homozygous hiPSC-NPCs may be useful for treating ischaemic stroke patients.

Multimodal Therapeutic Effects of Neural Precursor Cells Derived from Human-Induced Pluripotent Stem Cells through Episomal Plasmid-Based Reprogramming in a Rodent Model of Ischemic Stroke.

Stem cell therapy is a promising option for treating functional deficits in the stroke-damaged brain. Induced pluripotent stem cells (iPSCs) are attractive sources for cell therapy as they can be efficiently differentiated into neural lineages. Episomal plasmids (EPs) containing reprogramming factors can induce nonviral, integration-free iPSCs. Thus, iPSCs generated by an EP-based reprogramming technique (ep-iPSCs) have an advantage over gene-integrating iPSCs for clinical applications. However, there are few studies regarding the in vivo efficacy of ep-iPSCs. In this study, we investigated the therapeutic potential of intracerebral transplantation of neural precursor cells differentiated from ep-iPSCs (ep-iPSC-NPCs) in a rodent stroke model. The ep-iPSC-NPCs were transplanted intracerebrally in a peri-infarct area in a rodent stroke model. Rats transplanted with fibroblasts and vehicle were used as controls. The ep-iPSC-NPC-transplanted animals exhibited functional improvements in behavioral and electrophysiological tests. A small proportion of ep-iPSC-NPCs were detected up to 12 weeks after transplantation and were differentiated into both neuronal and glial lineages. In addition, transplanted cells promoted endogenous brain repair, presumably via increased subventricular zone neurogenesis, and reduced poststroke inflammation and glial scar formation. Taken together, these results strongly suggest that intracerebral transplantation of ep-iPSC-NPCs is a useful therapeutic option to treat clinical stroke through multimodal therapeutic mechanisms.

Triglyceride/HDL-Cholesterol Ratio as an Index of Intracranial Atherosclerosis in Nonstroke Individuals.

Background: Triglyceride (TG)/high-density lipoprotein cholesterol ratio (THR) is a marker of dyslipidemia, and high THR is associated with an increase in cardiovascular events. In the present study, whether THR was associated with various markers of cerebral vascular pathologies, atherosclerosis of major cerebral arteries, including large artery atherosclerosis (LAA) and cerebral small vessel disease (SVD), in neurologically healthy individuals was investigated. Methods: Vascular risk factors, brain magnetic resonance imaging (MRI) scans, and MR angiograms of 851 study subjects were evaluated. Findings of extracranial atherosclerosis (ECAS) and intracranial atherosclerosis (ICAS) were considered indices of LAA based on brain MR angiograms. The presence of silent lacunar infarct (SLI) and white matter hyperintensities (WMHs) were evaluated as indices of SVD based on brain MRIs. Results: Subjects with ICAS (odds ratio, 1.83; 95% confidence interval, 1.06-3.16; P = 0.03) were significantly more likely to have high THR tertile (THR > 2.06) than low THR tertile (THR < 1.37) after adjusting for cardiovascular risk factors. THR was higher in subjects with multiple ICAS lesions than in those with single ECAS or without ICAS lesions. Associations among THR tertiles in ECAS, SLI, and WMHs were not significant. Conclusion: In the present study, a positive association between high THR and the development of ICAS was observed in neurologically healthy participants.

Differential Impact of Plasma Homocysteine Levels on the Periventricular and Subcortical White Matter Hyperintensities on the Brain.

Background: The clinical significance of cerebral white matter hyperintensities (WMH) on brain magnetic resonance imaging (MRI) has recently increased, and recognized now as a risk factor for future stroke and dementia. High levels of plasma homocysteine (Hcyt) are associated with cerebral WMH. Recent studies suggest a different anatomy and physiology in the arteriolar system may be supplied to the periventricular and deep subcortical white matter. We hypothesize that plasma Hcyt levels have differing impacts on periventricular WMH (PVWMH) than on deep subcortical WMH (DSWMH). Methods: We evaluated plasma Hcyt levels from 937 neurologically healthy participants. The severity of PVWMH and DSWMH was evaluated by the use of a manual grading scale. Moderate to severe PVWMH and DSWMH levels were defined when the Fazekas score was two or three, respectively. Predominant PVWMH (pred-PVWMH) and predominant DSWMH (pred-DSWMH) were defined as having a difference of Fazekas score between PVWMH and DSWMH of two or more. Other confounding variables including age, sex, vascular risk factors, and estimated glomerular filtration rate (eGFR) were also analyzed. Results: Logistic regression revealed that, after adjusting for the confounding variables, PVWMH was associated with old age, hypertension, diabetes mellitus, low eGFR, and high plasma Hcyt levels. DSWMH was associated with old age, hypertension, and hypercholesterolemia but not with plasma Hcyt levels. Plasma Hcyt levels were associated with pred-PVWMH but not with pred-DSWMH. Conclusions: High plasma Hcyt levels are strongly associated with the development of PVWMH but not DSWMH. Our results suggest the possibility that different pathogeneses exist for PVWMH and DSWMH and that dysregulated Hcyt metabolism associated with the development of PVWMH.