Indoxyl sulfate contributes to colorectal cancer cell proliferation and increased EGFR expression by activating AhR and Akt.

Although patients with chronic kidney disease (CKD) have a higher risk of colorectal cancer (CRC) aggravation, the connection between these two diseases is not well understood. Recent studies have shown that both CKD and CRC aggravation are closely related to an increased abundance of indole-producing Fusobacterium nucleatum in the gut. The indole absorbed from the gut is eventually metabolized to indoxyl sulfate in the liver. Since indoxyl sulfate is involved not only in accelerating CKD progression but also in the initiation and development of its associated complications, the present study aimed to clarify whether indoxyl sulfate induces the proliferation of CRC cells. This study found that indoxyl sulfate induced the proliferation of CRC-derived HCT-116 cells by activating the aryl hydrocarbon receptor (AhR) and the proto-oncogene Akt. The AhR antagonist CH223191 and Akt inhibitor MK2206 suppressed indoxyl sulfate-induced proliferation of HCT-116 cells. We also found that indoxyl sulfate upregulated epidermal growth factor receptor (EGFR) expression, which is associated with poor prognosis of CRC, whereas CH223191 and MK2206 repressed EGFR expression. Furthermore, indoxyl sulfate increased the sensitivity of CRC cells to EGF by upregulating EGFR expression. These findings suggest that indoxyl sulfate may be an important link between CKD and CRC aggravation.

Effect of high-fat diet on cerebral pathological changes of cerebral small vessel disease in SHR/SP rats.

Cerebral small vessel diseases (CSVD) are neurological disorders associated with microvessels, manifested pathologically as white matter (WM) changes and cortical microbleeds, with hypertension as a risk factor. Additionally, a high-fat diet (HFD) can affect peripheral vessel health. Our study explored how HFD affects cerebral small vessels in normotensive WKY, hypertensive SHR, and SHR/SP rats. The MRI results revealed that HFD specifically increased WM hyperintensity in SHR/SP rats. Pathologically, it increased WM pallor and vacuolation in SHR and SHR/SP rats. Levels of blood-brain barrier (BBB) protein claudin 5 were decreased in SHR and SHR/SP compared to WKY, with HFD having minimal impact on these levels. Conversely, collagen IV levels remained consistent among the rat strains, which were increased by HFD. Consequently, HFD caused vessel leakage in all rat strains, particularly within the corpus callosum of SHR/SP rats. To understand the underlying mechanisms, we assessed the levels of hypoxia-inducible factor-1α (HIF-1α), Gp91-phox, and neuroinflammatory markers astrocytes, and microglia were increased in SHR and SHR/SP compared to WKY and were further elevated by HFD in all rat strains. Gp91-phox was also increased in SHR and SHR/SP compared to WKY, with HFD causing an increase in WKY but little effect in SHR and SHR/SP. In conclusion, our study demonstrates that HFD, in combined with hypertension, intensifies cerebral pathological alterations in CSVD rats. This exacerbation involves increased oxidative stress and HIF-1α in cerebral vessels, triggering neuroinflammation, vascular basement membrane remodeling, IgG leakage, and ultimately WM damage.

The Role of the Vascular System in Degenerative Diseases: Mechanisms and Implications.

Degenerative diseases, encompassing a wide range of conditions affecting various organ systems, pose significant challenges to global healthcare systems. This comprehensive review explores the intricate interplay between the vascular system and degenerative diseases, shedding light on the underlying mechanisms and profound implications for disease progression and management. The pivotal role of the vascular system in maintaining tissue homeostasis is highlighted, as it serves as the conduit for oxygen, nutrients, and immune cells to vital organs and tissues. Due to the vital role of the vascular system in maintaining homeostasis, its dysfunction, characterized by impaired blood flow, endothelial dysfunction, and vascular inflammation, emerges as a common denominator of degenerative diseases across multiple systems. In the nervous system, we explored the influence of vascular factors on neurodegenerative diseases such as Alzheimer's and Parkinson's, emphasizing the critical role of cerebral blood flow regulation and the blood-brain barrier. Within the kidney system, the intricate relationship between vascular health and chronic kidney disease is scrutinized, unraveling the mechanisms by which hypertension and other vascular factors contribute to renal dysfunction. Throughout this review, we emphasize the clinical significance of understanding vascular involvement in degenerative diseases and potential therapeutic interventions targeting vascular health, highlighting emerging treatments and prevention strategies. In conclusion, a profound appreciation of the role of the vascular system in degenerative diseases is essential for advancing our understanding of degenerative disease pathogenesis and developing innovative approaches for prevention and treatment. This review provides a comprehensive foundation for researchers, clinicians, and policymakers seeking to address the intricate relationship between vascular health and degenerative diseases in pursuit of improved patient outcomes and enhanced public health.

Correction: Shibly et al. Analysis of Cerebral Small Vessel Changes in AD Model Mice. Biomedicines 2023, 11, 50.

In the original publication [...].

Associations between dairy consumption and the physical function in Japanese community-dwelling older adults: The Shimane CoHRE study.

OBJECTIVES: We investigated sex differences in the associations between dairy consumption and the physical function among community-dwelling older adults. METHODS: Six hundred and fifty-six older adults (75.6 ± 6.4 years old) participated in this study. Dairy consumption (5-item Likert score) and the physical function (gait speed, handgrip strength, and skeletal muscle mass) were measured. The linear and quadratic associations between dairy consumption and the physical function measures were examined by a multiple linear regression analysis adjusted for covariates. RESULTS: Among women, an increased dairy consumption was significantly linearly associated with greater hand-grip strength and faster gait speed (both p<0.05) after adjusting for covariates. Among men, dairy consumption was not associated with the physical function measures. Dairy consumption was not associated with the muscle mass in either sex. CONCLUSIONS: Increased dairy consumption was associated with a superior physical function in older women.

Changes in masticatory performance and the new onset of diabetes mellitus in older adults: a 5-year longitudinal study.

Objective: Although masticatory performance is affected by age-related reduction in number of teeth (or treatment), the relationship between longitudinal changes in masticatory performance and diabetes mellitus is unknown. This longitudinal study investigated the association between changes in masticatory performance and new-onset diabetes mellitus among community-dwelling Japanese older adults. Methods: The data of 214 older adults living in Ohnan Town, Shimane, Japan, who participated in two surveys conducted between 2012 and 2017 were analyzed. Diabetes mellitus was defined as a hemoglobin A1c level ≥ 6.5% or self-reported diabetes mellitus. The masticatory performance (measured by number of gummy jelly pieces collected after chewing) was evaluated by dental hygienists. Masticatory performance was categorized into two groups (high or low) based on the median in each survey; further, four groups (Group A: remained consistently high, Group B: changed from low to high, Group C: remained consistently low, Group D: changed from high to low) were used to determine longitudinal changes in masticatory performance. Logistic regression was used to analyze the association between changes in masticatory performance and new-onset diabetes mellitus. Results: Overall, 10.3% of participants had diabetes mellitus at the follow-up survey. Multivariate analysis showed that Group D (odds ratio 8.69, 95% confidence interval 1.98-38.22) was positively associated with the development of diabetes mellitus compared with Group A after adjusting for sex, age, body mass index, alcohol consumption, physical activity, and eating speed. Conclusions: Deteriorating masticatory performance for 5 years may cause diabetes mellitus among older adults.

Alzheimer's Amyloid ß Peptide Induces Angiogenesis in an Alzheimer's Disease Model Mouse through Placental Growth Factor and Angiopoietin 2 Expressions.

Increased angiogenesis, especially the pathological type, has been documented in Alzheimer's disease (AD) brains, and it is considered to be activated due to a vascular dysfunction-mediated hypoxic condition. To understand the role of the amyloid ß (Aß) peptide in angiogenesis, we analyzed its effects on the brains of young APP transgenic AD model mice. Immunostaining results revealed that Aß was mainly localized intracellularly, with very few immunopositive vessels, and there was no extracellular deposition at this age. Solanum tuberosum lectin staining demonstrated that compared to their wild-type littermates, the vessel number was only increased in the cortex of J20 mice. CD105 staining also showed an increased number of new vessels in the cortex, some of which were partially positive for collagen4. Real-time PCR results demonstrated that placental growth factor (PlGF) and angiopoietin 2 (AngII) mRNA were increased in both the cortex and hippocampus of J20 mice compared to their wild-type littermates. However, vascular endothelial growth factor (VEGF) mRNA did not change. Immunofluorescence staining confirmed the increased expression of PlGF and AngII in the cortex of the J20 mice. Neuronal cells were positive for PlGF and AngII. Treatment of a neural stem cell line (NMW7) with synthetic Aß1-42 directly increased the expression of PlGF and AngII, at mRNA levels, and AngII at protein levels. Thus, these pilot data indicate that pathological angiogenesis exists in AD brains due to the direct effects of early Aß accumulation, suggesting that the Aß peptide regulates angiogenesis through PlGF and AngII expression.

Neither Trimethylamine-N-Oxide nor Trimethyllysine Is Associated with Atherosclerosis: A Cross-Sectional Study in Older Japanese Adults.

Recent evidence suggests that trimethylamine-N-oxide (TMAO), a metabolite of L-carnitine and choline, is linked to atherosclerosis and cardiovascular diseases. As TMAO content is very high in fish, we raised the following question: why do Japanese people, who consume lots of fish, show a low risk of atherosclerosis? To address this question, we investigated the effects of TMAO and other L-carnitine-related metabolites on carotid intima-media thickness (IMT). Participants were recruited from a small island and a mountainous region. Plasma L-carnitine, γ-butyrobetaine (γBB), TMAO, trimethyllysine (TML), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA) levels were measured using liquid or gas chromatography-mass spectrometry. Plasma L-carnitine concentration was higher in men than in women. TMAO and TML were significantly higher in the residents of the island than in the mountainous people. In multiple linear regression analyses in all participants, TML showed a significant inverse association with max-IMT and plaque score (PS), whereas TMAO did not show any associations. In women, L-carnitine was positively associated with max-IMT and PS. TMAO was correlated with both EPA and DHA levels, implying that fish is a major dietary source of TMAO in Japanese people. Our study found that plasma TMAO was not an apparent risk factor for atherosclerosis in elderly Japanese people, whereas a low level of TML might be a potential risk. L-carnitine may be a marker for atherosclerosis in women.

Serum Values of Cholesterol Absorption and Synthesis Biomarkers in Japanese Healthy Subjects: The CACHE Study HEALTHY Analysis.

AIM: Blood cholesterol absorption and synthesis biomarkers predict cardiovascular risk. This study aimed to determine the values of serum non-cholesterol sterol markers [lathosterol (Latho), campesterol (Campe), and sitosterol (Sito)] in healthy individuals and factors affecting these markers. METHODS: The CACHE Consortium compiled clinical data, including serum Latho (cholesterol synthesis marker), and Campe and Sito (cholesterol absorption markers), by a gas chromatography method in 2944 individuals. Healthy subjects were selected by excluding those with prior cardiovascular disease, diabetes mellitus, hypertension, chronic kidney disease, familial hypercholesterolemia, sitosterolemia, current smokers, those with low (＜17 kg/m2) or high (≥ 30 kg/m2) body mass index (BMI), and those with treatment for dyslipidemia or hyperuricemia. Nonlinear regression stratified by sex was used to examine the associations of cholesterol metabolism markers with age, BMI, and serum lipid levels. RESULTS: Of 479 individuals selected, 59.4% were female; the median age was 48 years in females and 50 years in males. The three markers showed positively skewed distributions, and sex differences were present. Age was associated positively with Latho, inversely with Campe, but not significantly with Sito. BMI was associated positively with Latho, but not significantly with Campe or Sito. High-density lipoprotein cholesterol (HDL-C) was positively associated with Campe and Sito, but not significantly with Latho. Non-HDL-C was positively associated with the three markers. CONCLUSION: Our study results in the healthy subjects help to interpret the non-cholesterol sterol markers for cardiovascular risk assessment in patients with cardiovascular risk factors.

Analysis of the time-dependent changes of phospholipids in the brain regions of a mouse model of Alzheimer's disease.

Phospholipid levels are reported to be decreased in Alzheimer's disease (AD). For a better understanding, we investigated the time-dependent changes of phospholipids species in a mouse model of AD. The levels of phospholipids in the hippocampus and prefrontal cortex of wild-type and APP-Tg (J20) mice were measured by LC-ESI-MS/MS. Compared to wild-type, total phosphatidylcholine (PC), phosphatidylethanolamine (PE), and lysophosphatidylcholine (LPC) were Increased at 3 months but decreased at 6 months in the cortex of J20 mice. Total lysophosphatidylethanolamine (LPE) was decreased both at 3 and 6 months. PC was decreased and LPC was increased at 6 months, resulting in an increased LPC/PC ratio in the hippocampus of J20 mice. At species levels, PCA analysis could discriminate wild-type and J20 based on PC and LPC distribution at 6 months. At 6 months, several highly abundant PC including PC (16:0/16:0), PC (16:0/18:0), PC (16:0/18:1), and PC (18:0/18:1) were decreased in the cortex and hippocampus of J20. Conversely, LPC species including LPC 16:0, LPC 18:1, and LPC 20:4 were increased especially in the hippocampal area. Increased activation of phospholipid-metabolizing enzyme cPLA2 was seen in the hippocampus and cortex of J20 mice at 9 months. On the other hand, ROS levels started to increase as early as 3 months. Compared to 3 months, ROS levels were higher at 6 months in J20 mice. Thus, we demonstrated here a time- and area-dependent alteration of phospholipid composition during the early stage of AD, which could be important in understanding the pathological process.

Changes in the Urinary Sodium-to-Potassium Ratio Are Associated with Blood Pressure Change in Older Japanese Adults: A 7-Year Longitudinal Study.

Studies on the association between sodium-to-potassium (Na/K) ratio changes and blood pressure (BP) changes among older adults are limited. This 7-year longitudinal study examined the association between Na/K ratio changes (evaluated using spot urine tests) and BP changes among older Japanese adults. Data were collected from 432 participants (mean age: 70.3±4.4; range: 65−84 years) in 2012 and 2019. Changes in BP and the Na/K ratio over 7 years were calculated by subtracting baseline values from values noted during a follow-up survey. The median systolic and diastolic BP (SBP) and (DBP) changes after 7 years were 4 (IQR, −7, 14) and −1 (IQR, −9, 5) mmHg, respectively. The median Na/K ratio was changed during the follow-up period by −0.2 (IQR, −1.3, 0.7). A generalized linear model indicated that Na/K ratio changes were positively associated with SBP (B = 2.03, p < 0.001) and DBP (B = 0.62, p = 0.021) changes. In the non-antihypertensive medication-using group, urinary Na/K ratio changes were associated with SBP and DBP changes (B = 2.39, p = 0.001; B = 0.99, p = 0.033). In the antihypertensive medication user group, urinary Na/K ratio changes were associated with SBP changes (B = 1.62, p = 0.015). We confirmed the association between changes in the Na/K ratio and changes in BP.

Rho-Kinase inhibition decreases focal cerebral ischemia-induced glial activation in rats.

Background: Rho-kinase inhibition in a rat middle cerebral artery occlusion (MCAO) model is reported to improve neurological functions and decrease infarction size. Objective: The objective of this study is to investigate the underlying mechanisms of such improvement by evaluating the effects of Rho-kinase inhibition on astrocytes and microglial accumulation and activation in this condition. Methods: Adult male Sprague-Dawley (SD) rats were used to generate the MCAO model, which received an I.P injection of a chemical Rho-kinase inhibitor (Fasudil- 5 mg/kg/day) or vehicle (PBS) for 2 and 4 days. Results: Fasudil treatment significantly decreased the stroke volumes and water content in the lesion areas, as revealed by MRI. Immunostaining and Western blotting results demonstrated that Fasudil significantly decreased the levels of Aquaporin-4, a water channel protein. The number of GFAP+ astrocytes and Iba-1+ macrophage/microglia was decreased in the lesion areas. Proinflammatory transcription factor NF-κB protein levels were decreased in the Fasudil group 2 days after MCAO. Also, proinflammatory mediators including TNF-α, IL-1ß, and iNOS levels were decreased. In vitro migration study using a human microglial cell line (HMO6) confirmed the inhibitory effects of Fasudil on the process. Fasudil also decreased combined IL-1ß and IFNγ-induced NF-κB nuclear translocation in HMO6. Moreover, Fasudil transiently decreased combined IL-1ß and IFNγ-induced iNOS, TNFα, and IL-1ß mRNA levels in HMO6. Conclusion: Our study demonstrates the inhibitory effects of Rho-kinase on NF-κB-mediated glial activation and cerebral edema, which might be a promising therapeutic target in acute cerebral ischemia conditions.

Associations between physical frailty and living arrangements in Japanese older adults living in a rural remote island: The Shimane CoHRE study.

Background: Living arrangements have been known to be associated with physical frailty. However, the prevalence of frailty and its risk factors in remote islands is not understood. We examined the association between living arrangements and objectively measured frailty among older adults living in a remote island of Japan. Methods: Among older people living in Okinoshima, 656 older adults (75.6 ± 6.4 years) were analyzed. Physical frailty (robust, prefrailty, or frailty) was assessed using the 5-item frailty phenotype (unintentional weight loss, self-reported exhaustion, weakness, slow walking speed, and low physical activity). Physical functions (muscle mass, gait speed, and grip strength) were measured objectively. Results: The prevalence of frailty and prefrailty was 6.6% and 43.8%, respectively. Living with a spouse resulted in a significantly lower prevalence of frailty (p < 0.001) compared with other living arrangements. All objectively measured physical functions among those who lived with a spouse were significantly superior to those who lived with family or alone (p < 0.001). Multinomial logistic regression showed that living alone was significantly associated with frailty (odds ratio [OR] 2.36, 95% confidence interval [CI] 1.07-5.24) and prefrailty (OR 1.75, 95% CI 1.14-2.69) after adjusting for all covariates. Conclusion: The prevalence of frailty on remote islands seemed similar to that in urban areas. Older people living in remote islands might be able to maintain their physical health. Furthermore, living alone may correlate with increased risks of frailty and prefrailty. Among elderly individuals on remote islands, living with a spouse might be desirable to prevent (pre)frailty.

Reduced masticatory performance and not using dentures are associated with hypertension in older adults with tooth loss: the Shimane CoHRE study.

The relationship between masticatory performance and hypertension has seldom been studied. The association between hypertension and tooth loss with/without denture use is unclear. This cross-sectional study examined the relationship between hypertension and masticatory performance and whether the relationship between hypertension and the number of teeth varied with denture use among older Japanese adults. Hypertension was defined as a systolic or diastolic blood pressure ≥140 or ≥90 mm Hg, respectively, or the use of antihypertensive medications. Masticatory performance was assessed using a chewable gummy jelly. The number of teeth and denture use were visually assessed by a trained dental hygienist. Poisson regression analyses were carried out to estimate the prevalence ratio and 95% confidence interval (CI) of hypertension with the number of teeth (≥28, 20-27, or 0-19 teeth), quartiles of masticatory performance (Q1: highest [reference] to Q4: lowest), and tooth loss with/without denture use. Hypertension was significantly associated with masticatory performance in participants in Q3 (prevalence ratio = 1.35; 95% CI = 1.07, 1.72) and Q4 (prevalence ratio = 1.31; 95% CI = 1.01, 1.69). There was no association between tooth loss and hypertension. Compared to participants with >28 teeth, the prevalence ratios and 95% CIs of those without dentures and those with 20-27 teeth were 1.33 (95% CI = 1.06, 1.68) and 1.69 (95% CI = 1.21, 2.37), respectively, in those with 0-19 teeth. Masticatory performance was inversely associated with hypertension, and tooth loss without the use of dentures was related to hypertension. Therefore, masticatory function may be important for preventing hypertension among older adults.

A Simple Flow Injection Analysis-Tandem Mass Spectrometry Method to Reduce False Positives of C5-Acylcarnitines Due to Pivaloylcarnitine Using Reference Ions.

Flow injection analysis−tandem mass spectrometry (FIA-TMS) has been applied in a first-tier test of newborn screening (NBS). Although isovalerylcarnitine (i-C5), which is a diagnostic indicator of isovaleric acidemia (IVA), is isobaric with pivaloylcarnitine (p-C5), 2-methylbutyrylcarnitine, and n-valerylcarnitine, these isomers cannot be distinguished by the FIA-TMS. There are many reports of false positives derived from p-C5 due to the use of pivalate-conjugated antibiotics. In this study, we developed a new FIA-TMS method to distinguish i-C5 and p-C5. We found that the intensity ratio of product ions for i-C5 and p-C5 was different in a certain range even under the same analytical conditions. The product ions with the most distinct differences in ionic intensity between the isomers and the collision energies that produce them were determined to be m/z 246.2 > 187.1 and −15 V, respectively. In addition to the quantification ion, a reference ion was defined, and the similarity of the i-C5 and p-C5 reference ion ratios (i-C5 score and p-C5 score, respectively) were used to estimate which isomer (i-C5 and p-C5) was responsible for elevated C5 acylcarnitine in dried blood spots (DBSs). As a result of analyses of 11 DBS samples derived from pivalate-conjugated antibiotics and four DBS samples from IVA patients using our method, it was found that our method was able to correctly determine the type of C5-acylcarnitine (i-C5 or p-C5) in the DBS samples. Implementation of this new FIA-TMS method into the current NBS protocol will allow for a reduction in false positives in IVA.

Combined association of oral and skeletal muscle health with type 2 diabetes mellitus among community-dwelling older adults in Japan: a cross-sectional study.

Objective: Although oral health and skeletal muscle status are known to be risk factors for type 2 diabetes mellitus (T2DM), there is limited information on their combined effects among community-dwelling older adults. The purpose of this study was to investigate the association between oral health and skeletal muscle status among older adults with T2DM in Japan. Participants and Methods: This cross-sectional study included data from individuals aged ≥60 years. T2DM was defined as a glycosylated hemoglobin A1c level ≥48 mmol/mol (≥6.5%) or the use of hypoglycemic agents. For oral health status, dental hygienists assessed the number of teeth (NT) and masticatory function (MF). Skeletal muscle status was assessed using skeletal muscle mass index (SMI) and handgrip strength (HGS). Logistic regression analysis examined T2DM in nine-category combinations of oral health status (each of the three categories in NT and MF) and skeletal status (each of the three categories in SMI and HGS). Results: T2DM was prevalent in 83 participants (16.4%) and was significantly associated with low NT and SMI (odds ratio [OR] = 5.93, 95% confidence interval [CI]: 1.37-25.73) and low MF and SMI (OR = 4.48, 95% CI: 1.23-16.35) compared to high NT and SMI and high MF and SMI, respectively. Conclusion: Our findings indicate that low muscle mass with tooth loss or masticatory dysfunction is associated with T2DM among community-dwelling older adults. This suggests that maintaining oral health and muscle mass may be an effective strategy for the prevention of T2DM.

iTRAQ-Based Proteomic Analysis of APP Transgenic Mouse Urine Exosomes.

Alzheimer's disease (AD) is a common dementia disease in the elderly. To get a better understanding of the pathophysiology, we performed a proteomic analysis of the urine exosomes (U-exo) in AD model mice (J20). The polymer precipitation method was used to isolate U-exo from the urine of 3-month-old J20 and wild-type (WT) mice. Neuron-derived exosome (N-exo) was isolated from U-exo by immunoprecipitation. iTRAQ-based MALDI TOF MS/MS was used for proteomic analysis. The results showed that compared to WT, the levels of 61 and 92 proteins were increased in the J20 U-exo and N-exo, respectively. Gene ontology enrichment analysis demonstrated that the sphingolipid catabolic process, ceramide catabolic process, membrane lipid catabolic process, Aß clearance, and Aß metabolic process were highly enriched in U-exo and N-exo. Among these, Asah1 was shown to be the key protein in lipid metabolism, and clusterin, ApoE, neprilysin, and ACE were related to Aß metabolism and clearance. Furthermore, protein-protein interaction analysis identified four protein complexes where clusterin and ApoE participated as partner proteins. Thus, J20 U-exo and N-exo contain proteins related to lipid- and Aß-metabolism in the early stages of AD, providing a new insight into the underlying pathological mechanism of early AD.

Analysis of Cerebral Small Vessel Changes in AD Model Mice.

Amyloid ß (Aß) peptide is deposited in the brains of sporadic Alzheimer's disease (AD) due to impaired vessel-dependent clearance. To understand the mechanisms, we investigated time-dependent cerebrovascular changes in AD model mice. Cerebrovascular and other pathological changes were analyzed in AD model mice (J20 strain) aging from 2 to 9 months by immunostaining. At 2 months, Aß was only intraneuronal, whereas vessels were positive from 3 months in J20 mice. Compared to wild-type (WT), vessel density was increased at 2 months but decreased at 9 months in J20 mice, claudin-5 levels were decreased, and vascular endothelial growth factor (VEGF) levels were increased in the cortex and hippocampus of J20 mice brain at all time points. Albumin extravasation was evident from 3 months in J20 brains. Collagen 4 was increased at 2 and 3 months. Aquaporin 4 was spread beyond the vessels starting from 3 months in J20, which was restricted around the vessel in wild-type mice. In conclusion, the study showed that an early decrease in claudin-5 was associated with VEGF expression, indicating dysfunction of the blood-brain barrier. Decreased claudin-5 might cause the leakage of blood constituents into the parenchyma that alters astrocyte polarity and its functions.