Candesartan prevents arteriopathy progression in cerebral autosomal recessive arteriopathy with subcortical infarcts and leukoencephalopathy model.

Cerebral small vessel disease (CSVD) causes dementia and gait disturbance due to arteriopathy. Cerebral autosomal recessive arteriopathy with subcortical infarcts and leukoencephalopathy (CARASIL) is a hereditary form of CSVD caused by loss of high-temperature requirement A1 (HTRA1) serine protease activity. In CARASIL, arteriopathy causes intimal thickening, smooth muscle cell (SMC) degeneration, elastic lamina splitting, and vasodilation. The molecular mechanisms were proposed to involve the accumulation of matrisome proteins as substrates or abnormalities in transforming growth factor ß (TGF-ß) signaling. Here, we show that HTRA1-/- mice exhibited features of CARASIL-associated arteriopathy: intimal thickening, abnormal elastic lamina, and vasodilation. In addition, the mice exhibited reduced distensibility of the cerebral arteries and blood flow in the cerebral cortex. In the thickened intima, matrisome proteins, including the hub protein fibronectin (FN) and latent TGF-ß binding protein 4 (LTBP-4), which are substrates of HTRA1, accumulated. Candesartan treatment alleviated matrisome protein accumulation and normalized the vascular distensibility and cerebral blood flow. Furthermore, candesartan reduced the mRNA expression of Fn1, Ltbp-4, and Adamtsl2, which are involved in forming the extracellular matrix network. Our results indicate that these accumulated matrisome proteins may be potential therapeutic targets for arteriopathy in CARASIL.

Excessive Production of Transforming Growth Factor ß1 Causes Mural Cell Depletion From Cerebral Small Vessels.

It is increasingly becoming apparent that cerebrovascular dysfunction contributes to the pathogenic processes involved in vascular dementia, Alzheimer's disease, and other neurodegenerative disorders. Under these pathologic conditions, the degeneration of cerebral blood vessels is frequently accompanied by a loss of mural cells from the vascular walls. Vascular mural cells play pivotal roles in cerebrovascular functions, such as regulation of cerebral blood flow and maintenance of the blood-brain barrier (BBB). Therefore, cerebrovascular mural cell impairment is involved in the pathophysiology of vascular-related encephalopathies, and protecting these cells is essential for maintaining brain health. However, our understanding of the molecular mechanism underlying mural cell abnormalities is incomplete. Several reports have indicated that dysregulated transforming growth factor ß (TGFß) signaling is involved in the development of cerebral arteriopathies. These studies have specifically suggested the involvement of TGFß overproduction. Although cerebrovascular toxicity via vascular fibrosis by extracellular matrix accumulation or amyloid deposition is known to occur with enhanced TGFß production, whether increased TGFß results in the degeneration of vascular mural cells in vivo remains unknown. Here, we demonstrated that chronic TGFß1 overproduction causes a dropout of mural cells and reduces their coverage on cerebral vessels in both smooth muscle cells and pericytes. Mural cell degeneration was also accompanied by vascular luminal dilation. TGFß1 overproduction in astrocytes significantly increased TGFß1 content in the cerebrospinal fluid (CSF) and increased TGFß signaling-regulated gene expression in both pial arteries and brain capillaries. These results indicate that TGFß is an important effector that mediates mural cell abnormalities under pathological conditions related to cerebral arteriopathies.

Spectral domain optical coherence tomography and fundus autofluorescence findings in cytomegalovirus retinitis in HIV-infected patients.

PURPOSE: To assess the usefulness of spectral domain optical coherence tomography (SD-OCT) and fundus autofluorescence (FAF) findings in cytomegalovirus (CMV) retinitis. STUDY DESIGN: Observational case series. METHODS: Thirteen eyes of 11 human immunodeficiency virus (HIV)-positive patients with CMV retinitis underwent full ophthalmologic examinations, SD-OCT, and 4 eyes of 4 patients underwent FAF. FAF images included short-wavelength autofluorescence (SW-AF) and near-infrared autofluorescence (IR-AF). CMV retinitis was classified into proposed categories of acute, subacute, remission, and recurrent; the acute stage was further subdivided into initial, early, and late stages. RESULTS: In the initial stage, vertical structural disruption of all retinal layers was observed by SD-OCT, and FAF showed hyperautofluorescence on SW-AF and hypoautofluorescence on IR-AF. In the early stage, SD-OCT showed significant retinal thickening; cells and debris from the retinal surface to the vitreous; enlarged vessels with/without thickened vessel walls; and highly complicated serous retinal detachment. In the late to subacute stage, features observed included rhegmatogenous retinal detachment with shrinking posterior hyaloid membrane and waving from the ellipsoid zone to the retinal pigment epithelium. In remission, FAF findings were hypoautofluorescence on SW-AF and hyperautofluorescence on IR-AF. CONCLUSION: Although the number of examined eyes was limited, SD-OCT and FAF provide new information in various stages of CMV retinitis in patients with HIV infection that is not obtainable by conventional examination and which may be of great benefit when screening for the initial stage of CMV retinitis.

Distinct molecular mechanisms of HTRA1 mutants in manifesting heterozygotes with CARASIL.

OBJECTIVE: To elucidate the molecular mechanism of mutant HTRA1-dependent cerebral small vessel disease in heterozygous individuals. METHODS: We recruited 113 unrelated index patients with clinically diagnosed cerebral small vessel disease. The coding sequences of the HTRA1 gene were analyzed. We evaluated HTRA1 protease activities using casein assays and oligomeric HTRA1 formation using gel filtration chromatography. RESULTS: We found 4 heterozygous missense mutations in the HTRA1 gene (p.G283E, p.P285L, p.R302Q, and p.T319I) in 6 patients from 113 unrelated index patients and in 2 siblings in 2 unrelated families with p.R302Q. The mean age at cognitive impairment onset was 51.1 years. Spondylosis deformans was observed in all cases, whereas alopecia was observed in 3 cases; an autopsied case with p.G283E showed arteriopathy in their cerebral small arteries. These mutant HTRA1s showed markedly decreased protease activities and inhibited wild-type HTRA1 activity, whereas 2 of 3 mutant HTRA1s reported in cerebral autosomal-recessive arteriopathy with subcortical infarcts and leukoencephalopathy (CARASIL) (A252T and V297M) did not inhibit wild-type HTRA1 activity. Wild-type HTRA1 forms trimers; however, G283E and T319I HTRA1, observed in manifesting heterozygotes, did not form trimers. P285L and R302Q HTRA1s formed trimers, but their mutations were located in domains that are important for trimer-associated HTRA1 activation; in contrast, A252T and V297M HTRA1s, which have been observed in CARASIL, also formed trimers but had mutations outside the domains important for trimer-associated HTRA1 activation. CONCLUSIONS: The mutant HTRA1s observed in manifesting heterozygotes might result in an impaired HTRA1 activation cascade of HTRA1 or be unable to form stable trimers.

Characteristic features and progression of abnormalities on MRI for CARASIL.

OBJECTIVES: The objective of this study was to clarify the characteristic brain MRI findings for genetically diagnosed CARASIL (cerebral autosomal recessive arteriopathy with subcortical infarcts and leukoencephalopathy). METHODS: Seven patients with CARASIL carrying HTRA1 mutations (representing 6 Japanese families) were included in this study. Eighteen brain MRIs were reviewed and evaluated with a new rating scale based on scoring for abnormal hyperintense lesions and atrophy. RESULTS: At the last follow-up MRI, all patients had hyperintense lesions on T2-weighted images of the frontal white matter, anterior temporal lobe, external capsules, and thalami. Patients with longer time from the onset of cognitive impairment had higher MRI severity score. The atrophy advanced, followed by white matter lesion progression. During the early stage, hyperintense lesions were observed in the frontal white matter, external capsule, and pons. During the late stage, the arc-shaped hyperintense lesion from the pons to the middle cerebellar peduncles, which we designated the "arc sign," became evident. The arc sign was a characteristic finding for CARASIL in the advanced stage. CONCLUSIONS: These characteristic MRI findings for CARASIL are useful for selecting patients for genetic testing. The rating scale correlates well with disease duration and might be useful for assessing disease progression.

[Multimodal imaging of a case of acute syphilitic posterior placoid chorioretinitis].

BACKGROUND: Acute syphilitic posterior placoid chorioretinitis (ASPPC) is a rare manifestation of ocular syphilis. We report on multimodal imaging including ophthalmoscopy, spectral-domain optical coherence tomography (SD-OCT), fluorescein angiography (FA) and indocyanine green angiography (IA) of a case diagnosed with ASPPC. CASE: A 45-year-old man who was positive for human immunodeficiency virus presented with a 2-week history of visual loss in the right eye. CLINICAL FINDINGS: Ophthalmoscopy showed a unilateral yellowish lesion involving the macula. SD-OCT revealed absence of the photoreceptor inner segment ellipsoid as well as an absent external limiting membrane, and nodular elevations of the retinal pigment epithelium layer at the macula. Late IA demonstrated punctate hypofluorescent dots in diffuse hyperfluorescent area corresponding to the macular lesion. Serologic tests were positive for syphilis and the patient was treated with intravenous penicillin G. Visual acuity improved with treatment from 20/100 to 20/16 and the retinal appearance returned to normal. There was completely restored stratification of the outer retina after therapy. CONCLUSION: In the present case of ASPPC, the SD-OCT imaging demonstrated characteristic abnormalities including RPE nodularity which showed hypofluorescent dots on late IA.

Sporadic ALS with compound heterozygous mutations in the SQSTM1 gene.

Accumulating evidence suggests that heterozygous mutations in the SQSTM1 gene, which encodes p62 protein, are associated with amyotrophic lateral sclerosis (ALS). Here, we report a Japanese patient with sporadic, late-onset ALS who harbored compound heterozygous SQSTM1 mutations (p.[Val90Met];[Val153Ile]). Autopsy examination revealed that although TDP-43 pathology was rather widespread, the selective occurrence of p62-positive/TDP-43-negative cytoplasmic inclusions in the lower motor neurons (LMNs) was a characteristic feature. No Bunina bodies were found. Ultrastructurally, p62-positive cytoplasmic inclusions observed in the spinal anterior horn cells were composed of aggregates of ribosome-like granules and intermingled bundles of filamentous structures. Another feature of interest was concomitant Lewy body pathology. The occurrence of distinct p62 pathology in the LMNs in this patient indicates the pathogenic role of SQSTM1 mutations in the development of a subset of ALS.

Hyperphosphorylation of Tau induced by naturally secreted amyloid-ß at nanomolar concentrations is modulated by insulin-dependent Akt-GSK3ß signaling pathway.

Alzheimer disease (AD) is neuropathologically characterized by the formation of senile plaques from amyloid-ß (Aß) and neurofibrillary tangles composed of phosphorylated Tau. Although there is growing evidence for the pathogenic role of soluble Aß species in AD, the major question of how Aß induces hyperphosphorylation of Tau remains unanswered. To address this question, we here developed a novel cell coculture system to assess the effect of extracellular Aß at physiologically relevant levels naturally secreted from donor cells on the phosphorylation of Tau in recipient cells. Using this assay, we demonstrated that physiologically relevant levels of secreted Aß are sufficient to cause hyperphosphorylation of Tau in recipient N2a cells expressing human Tau and in primary culture neurons. This hyperphosphorylation of Tau is inhibited by blocking Aß production in donor cells. The expression of familial AD-linked PSEN1 mutants and APP ΔE693 mutant that induce the production of oligomeric Aß in donor cells results in a similar hyperphosphorylation of Tau in recipient cells. The mechanism underlying the Aß-induced Tau hyperphosphorylation is mediated by the impaired insulin signal transduction because we demonstrated that the phosphorylation of Akt and GSK3ß upon insulin stimulation is less activated under this condition. Treating cells with the insulin-sensitizing drug rosiglitazone, a peroxisome proliferator-activated receptor γ agonist, attenuates the Aß-dependent hyperphosphorylation of Tau. These findings suggest that the disturbed insulin signaling cascade may be implicated in the pathways through which soluble Aß induces Tau phosphorylation and further support the notion that correcting insulin signal dysregulation in AD may offer a potential therapeutic approach.

Combination of abducens nerve palsy and ipsilateral postganglionic Horner syndrome as an initial manifestation of uterine cervical cancer.

A 74-year-old woman presented with abducens nerve palsy, postganglionic Horner syndrome and sensory disturbance in the territory of the ophthalmic nerve on the left side. Cranial magnetic resonance imaging demonstrated a gadolinium-enhanced lesion within the left cavernous sinus. Thereafter, uterine cervical cancer was detected as the origin of this intra-cavernous sinus metastasis. We emphasize that the combination of abducens nerve palsy and ipsilateral postganglionic Horner syndrome may indicate a lesion located within the posterior portion of the cavernous sinus or in its vicinity. Moreover, this is the first reported case of uterine cervical cancer with intra-cavernous sinus metastasis.

Quantitative atlas of membrane transporter proteins: development and application of a highly sensitive simultaneous LC/MS/MS method combined with novel in-silico peptide selection criteria.

PURPOSE: To develop an absolute quantification method for membrane proteins, and to construct a quantitative atlas of membrane transporter proteins in the blood-brain barrier, liver and kidney of mouse. METHODS: Mouse tissues were digested with trypsin, and mixed with stable isotope labeled-peptide as a quantitative standard. The amounts of transporter proteins were simultaneously determined by liquid chromatography-tandem mass spectrometer (LC/MS/MS). RESULTS: The target proteins were digested in-silico, and target peptides for analysis were chosen on the basis of the selection criteria. All of the peptides selected exhibited a detection limit of 10 fmol and linearity over at least two orders of magnitude in the calibration curve for LC/MS/MS analysis. The method was applied to obtain the expression levels of 34 transporters in liver, kidney and blood-brain barrier of mouse. The quantitative values of transporter proteins showed an excellent correlation with the values obtained with existing methods using antibodies or binding molecules. CONCLUSION: A sensitive and simultaneous quantification method was developed for membrane proteins. By using this method, we constructed a quantitative atlas of membrane transporter proteins at the blood-brain barrier, liver and kidney in mouse. This technology is expected to have major implications for various fields of biomedical science.