Immunoreactivity and a new staining method of monocarboxylate transporter 1 located in endothelial cells of cerebral vessels of human brain in distinguishing cerebral venules from arterioles

Abstract

Distinguishing brain venules from arterioles with arteriolosclerosis is less reliable using traditional staining methods. We aimed to immunohistochemically assess the monocarboxylate transporter 1 (MCT1), a specific marker of venous endothelium found in rodent studies, in different caliber vessels in human brains. Both largeand small-caliber cerebral vessels were dissected from four autopsy donors. Immunoreactivity for MCT1 was examined in all autopsied human brain tissues, and then each vessel was identified by neuropathologists using hematoxylin and eosin stain, the Verhoeff’s Van Gieson stain, immunohistochemical stain with antibodies for α-smooth muscle actin and MCT1 in sequence. A total of 61 cerebral vessels, including 29 arteries and 32 veins were assessed. Immunoreactivity for MCT1 was observed in the endothelial cells of various caliber veins as well as the capillaries, whereas that was immunenegative in the endothelium of arteries. The different labeling patterns for MCT1 could aid in distinguishing various caliber veins from arteries, whereas assessment using the vessel shape, the internal elastic lamina, and the pattern of smooth muscle fibers failed to make the distinction between small-caliber veins and sclerotic arterioles. In conclusion, MCT1 immunohistochemical staining is a sensitive and reliable method to distinguish cerebral veins from arteries.

Arteriolosclerosis differs from venular collagenosis in relation to cerebrovascular parenchymal damages: an autopsy-based study

“…6 10 Anti-MCT1 immunohistochemical staining, a sensitive and reliable method for distinguishing cerebral venules from arterioles, combined with anti-αSMA, was used in our study to overcome these problems. 8 We found that although the severity of arteriolosclerosis significantly increased with venular collagenosis across the brain regions, an inverse severity distribution was found in 20%-30% of cases. These data suggest that, despite some shared risk factors, the mechanisms underlying the development arteriolosclerosis and venous collagenosis might differ.…”

“… 6 10 Anti-MCT1 immunohistochemical staining, a sensitive and reliable method for distinguishing cerebral venules from arterioles, combined with anti-αSMA, was used in our study to overcome these problems. 8 …”

“…Based on our previous report, arterioles were identified as negative immunolabeled for MCT1 and generally positive immunolabeled for αSMA. 8 According to the rating scheme for cerebrovascular lesions, arterioles with a score of 2 or 3 were assessed as arteriosclerotic arterioles in our study, following which the percentage of arteriolosclerosis was calculated. 9 Similarly, for venular collagenosis evaluation, vessels ranging from 10 to 300 µm in diameter, which showed intense labelling for MCT1 in the endothelium with a thin anti-αSMA-stained pattern in the media tunica, were considered as venules, 8 and those with severe stenosis (the lumen occupying more than 50% of the vessel diameter) or occlusion by thick Open access collagenous walls were assessed as venular collagenosis (figure 1) according to previous scores by Moody et al 4 The ratio of the number of collagenised venules to the total number of venules in the visual field was calculated and expressed as a percentage of venular collagenosis.…”

“…Immunohistochemistry was performed using antibodies against alpha-smooth muscular actin (αSMA, Cat# 19245, RRID: AB_2734735, rabbit monoclonal antibody, diluted 1:400; Cell Signaling Technology) and monocarboxylate transporter 1 (MCT1, Cat# 20 139-1-AP, RRID: AB_2878645, rabbit polyclonal antibody, diluted 1:800; Proteintech)to distinguish between venules and arterioles(details of primary antibodies and antigen retrieval protocols for immunohistochemistry are presented in online supplemental table 2 ). 8 Immunopositivity was detected using the VECTASTAIN ABC-HRP Kit (rabbit IgG, PK-4001; Vector Laboratories, Burlingame, California, USA) with 3,3 diaminobenzidine as a chromogen and hematoxylin as the counterstain. All histologically and immunohistochemically stained sections were subsequently dehydrated using a series of alcohols, cleared, and mounted in neutral balsam (ZLI-9555, ZSGB-BIO, Beijing, China).…”

Histopathological Analysis of Cerebral Microvasculature Pathology with Alzheimer’s Disease Neuropathological Changes: An Autopsy Study

“…To evaluate the severity of arteriolosclerosis and venular collagenosis, 2400 µm × 3800 µm single images were captured randomly in anti-αSMA and anti-MCT1 immunostained sections to differentiate between arterioles and venules, and the corresponding visual elds on the delineated images were subsequently captured in adjacent Masson's trichrome sections to assess the severity. For arteriolosclerosis evaluation, based on our previous report [25], vessels from 10 to 200 µm in diameter that showed negative immunolabeling for MCT1 and positive immunolabeling for αSMA were considered arterioles. According to the Vascular Cognitive Impairment Neuropathology Guidelines (VCING)[26], arteriolosclerosis was graded according to severity using histological semiquantitative scales ranging from 0 to 3 (0 = normal, 1 = mild changes, 2 = moderate changes, and 3 = severe changes).…”