IL-18 (Interleukin-18) Produced by Renal Tubular Epithelial Cells Promotes Renal Inflammation and Injury During Deoxycorticosterone/Salt-Induced Hypertension in Mice.

IL-18 (interleukin-18) is elevated in hypertensive patients, but its contribution to high blood pressure and end-organ damage is unknown. We examined the role of IL-18 in the development of renal inflammation and injury in a mouse model of low-renin hypertension. Hypertension was induced in male C57BL6/J (WT) and IL-18−/− mice by uninephrectomy, deoxycorticosterone acetate (2.4 mg/d, s.c.) and 0.9% drinking saline (1K/DOCA/salt). Normotensive controls received uninephrectomy and placebo (1K/placebo). Blood pressure was measured via tail cuff or radiotelemetry. After 21 days, kidneys were harvested for (immuno)histochemical, quantitative-PCR and flow cytometric analyses of fibrosis, inflammation, and immune cell infiltration. 1K/DOCA/salt-treated WT mice developed hypertension, renal fibrosis, upregulation of proinflammatory genes, and accumulation of CD3+ T cells in the kidneys. They also displayed increased expression of IL-18 on tubular epithelial cells. IL-18−/− mice were profoundly protected from hypertension, renal fibrosis, and inflammation. Bone marrow transplantation between WT and IL-18−/− mice revealed that IL-18-deficiency in non-bone marrow-derived cells alone afforded equivalent protection against hypertension and renal injury as global IL-18 deficiency. IL-18 receptor subunits­interleukin-18 receptor 1 and IL-18R accessory protein­were upregulated in kidneys of 1K/DOCA/salt-treated WT mice and localized to T cells and tubular epithelial cells. T cells from kidneys of 1K/DOCA/salt-treated mice produced interferon-γ upon ex vivo stimulation with IL-18, whereas those from 1K/placebo mice did not. In conclusion, IL-18 production by tubular epithelial cells contributes to elevated blood pressure, renal inflammation, and fibrosis in 1K/DOCA/salt-treated mice, highlighting it as a promising therapeutic target for hypertension and kidney disease.

Aldosterone-induced hypertension is sex-dependent, mediated by T cells and sensitive to GPER activation.

AIMS: The G protein-coupled estrogen receptor 1 (GPER) may modulate some effects of aldosterone. In addition, G-1 (a GPER agonist) can lower blood pressure (BP) and promote T cell-mediated anti-inflammatory responses. This study aimed to test the effects of G-1 and G-15 (a GPER antagonist) on aldosterone-induced hypertension in mice and to examine the cellular mechanisms involved. METHODS AND RESULTS: C57Bl/6 (wild-type, WT), RAG1-deficient and GPER-deficient mice were infused with vehicle, aldosterone (0.72 mg/kg/day S.C. plus 0.9% NaCl for drinking) ± G-1 (0.03 mg/kg/day S.C.) ± G-15 (0.3 mg/kg/day S.C.) for 14 days. G-1 attenuated aldosterone-induced hypertension in male WT but not male GPER-deficient mice. G-15 alone did not alter hypertension but it prevented the anti-hypertensive effect of G-1. In intact female WT mice, aldosterone-induced hypertension was markedly delayed and suppressed compared with responses in males, with BP remaining unchanged until after Day 7. In contrast, co-administration of aldosterone and G-15 fully increased BP within 7 days in WT females. Similarly, aldosterone robustly increased BP by Day 7 in ovariectomized WT females, and in both sexes of GPER-deficient mice. Whereas aldosterone had virtually no effect on BP in RAG1-deficient mice, adoptive transfer of T cells from male WT or male GPER-deficient mice into male RAG1-deficient mice restored the pressor response to aldosterone. This pressor effect could be attenuated by G-1 in RAG1-deficient mice that were reconstituted with either WT or GPER-deficient T cells, suggesting that G-1 does not act via T cells to lower BP. CONCLUSION: Our findings indicate that although aldosterone-induced hypertension is largely mediated by T cells, it can be attenuated by activation of GPER on non-T cells, which accounts for the sex difference in sensitivity to the pressor effect.

Pharmacological inhibition of the NLRP3 inflammasome reduces blood pressure, renal damage, and dysfunction in salt-sensitive hypertension.

AIMS: Renal inflammation, leading to fibrosis and impaired function is a major contributor to the development of hypertension. The NLRP3 inflammasome mediates inflammation in several chronic diseases by processing the cytokines pro-interleukin (IL)-1ß and pro-IL-18. In this study, we investigated whether MCC950, a recently-identified inhibitor of NLRP3 activity, reduces blood pressure (BP), renal inflammation, fibrosis and dysfunction in mice with established hypertension. METHODS AND RESULTS: C57BL6/J mice were made hypertensive by uninephrectomy and treatment with deoxycorticosterone acetate (2.4 mg/day, s.c.) and 0.9% NaCl in the drinking water (1K/DOCA/salt). Normotensive controls were uninephrectomized and received normal drinking water. Ten days later, mice were treated with MCC950 (10 mg/kg/day, s.c.) or vehicle (saline, s.c.) for up to 25 days. BP was monitored by tail-cuff or radiotelemetry; renal function by biochemical analysis of 24-h urine collections; and kidney inflammation/pathology was assessed by real-time PCR for inflammatory gene expression, flow cytometry for leucocyte influx, and Picrosirius red histology for collagen. Over the 10 days post-surgery, 1K/DOCA/salt-treated mice became hypertensive, developed impaired renal function, and displayed elevated renal levels of inflammatory markers, collagen and immune cells. MCC950 treatment from day 10 attenuated 1K/DOCA/salt-induced increases in renal expression of inflammasome subunits (NLRP3, ASC, pro-caspase-1) and inflammatory/injury markers (pro-IL-18, pro-IL-1ß, IL-17A, TNF-α, osteopontin, ICAM-1, VCAM-1, CCL2, vimentin), each by 25-40%. MCC950 reduced interstitial collagen and accumulation of certain leucocyte subsets in kidneys of 1K/DOCA/salt-treated mice, including CD206+ (M2-like) macrophages and interferon-gamma-producing T cells. Finally, MCC950 partially reversed 1K/DOCA/salt-induced elevations in BP, urine output, osmolality, [Na+], and albuminuria (each by 20-25%). None of the above parameters were altered by MCC950 in normotensive mice. CONCLUSION: MCC950 was effective at reducing BP and limiting renal inflammation, fibrosis and dysfunction in mice with established hypertension. This study provides proof-of-concept that pharmacological inhibition of the NLRP3 inflammasome is a viable anti-hypertensive strategy.

Age-Related Changes in the Chorioretinal Junction: An Immunohistochemical Study.

The chorioretinal junction comprises the retinal pigment epithelium, Bruch's membrane (BM), and adjacent choroidal capillaries. Its significance lies in its ability to support the retina mechanically and metabolically. The aim of this cross-sectional study was to record the senescent changes affecting all the constituents of the chorioretinal junction in 40 histological specimens across the whole spectrum of the adult age range. This study included light microscopy, with hematoxylin and eosin and PAS stains, and fluorescent microscopy. Immunohistochemistry was done using antibodies against neurofilament, synaptophysin, S-100, and collagen IV. The descriptive microanatomy was corroborated by morphometry. The amount of melanin and lipofuscin granule and drusens were noted. The ratio of thickness of BM to capillary diameter reduced from 1:6 or less in the 2nd decade to 1:3 in the 10th decade. Complete hyalinization of intercapillary pillars was seen in the 10th decade. The accumulation of lipofuscin with age was documented with the diminution in the size of epithelial cells. The subepithelial accumulation of drusen was first noted in the specimen from the late 60s. We have described all senescent changes in the chorioretinal junction chronologically. Similar changes are found in a more pronounced form in age-related macular degeneration. These data might serve as a reference baseline for clinicians and pathologists.

Human retinal senescence measured histologically: as an aid to clinical diagnosis by ocular coherence tomography

Spectral domain optical coherence tomography (SD-OCT) has become an established diagnostic tool for the clinical assessment of retinal pathology and its progression, in OPD setting. The aim of our study was to do in vitro quantification of relevant retinal layers to collect baseline data at different age groups, against which OCT findings can be interpreted. Thirty eyeballs (20-99 years) were used to study the retinal nerve fibre layer (RNFL), ganglion cells and inner plexiform layer (GC+IPL) and outer nuclear layer (ONL) thickness using V-Test software, 198035 on the H&E stained histological specimens. Mean thickness of these retinal layers was studied. To estimate the decrease from the optimal stage, absolute percentage decline (APD) was calculated for each decade. The mean thickness of RNFL at was 77.8 µm, 77.1 µm, 73.6 µm, 70.6 µm, 69.2µm, 54.1µm, 36.5µm 26.8µm from 3rd to 10th decade respectively. Significant APD of 30% was evident between 7th and 8th decades. APD graphs for RNFL and GC+IPL were almost parallel to each other. The absolute percentage decline (APD) for the thickness of ONL was 0.2%, 4.6%, 13%, 35%, 60%, 62% and 64% for 4th, 5th, 6th, 7th, 8th, 9th and 10th decade respectively. This study has provided normative base line histological data for ready reference. Decade wise changes in thickness of different layers can be used by ophthalmologist to differentiate senescent from pathological changes and to monitor progression of disease

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Vascularity of human atrioventricular valves: a myth or fact?

BACKGROUND: Knowledge of heart valve vascularity is an important factor for understanding the valvular pathology and to develop tissue-engineered valves for repair procedures. Some investigators believe that blood vessels may exist in normal human heart valves whereas some recent publications have proposed that the presence of blood vessels in the valves is secondary to inflammation. METHODS: Tissues from 60 normal formalin-fixed human hearts were examined microscopically for type, location, and number of vessels in atrioventricular valves. The age of the patient ranged from 10 to 70 years, and an attempt was made to study the age-related morphologic changes in atrioventricular valves. RESULTS: Of the 60 tricuspid and 60 mitral valves examined, 12 tricuspid (20%) and 14 mitral (23.33%) valves were found to have vessels without the presence of an inflammatory process. In tricuspid valves the vessels were observed mainly in the fibrosa layer with a range of 1 to 4 vessels, whereas in mitral valves the vessels were situated mainly in the spongiosa layer with a range of 1 to 2 vessels. The maximum vascularity was seen in the fourth decade of life, in which the vessels were found in 40% of both tricuspid and mitral valves. The mean transverse diameter of these vessels was 0.23 ± 0.18 mm, with a range of 0.06 to 0.79 mm in tricuspid valves, whereas it was 0.15 ± 0.08 mm, with a range of 0.04 to 0.4 mm in mitral valves. The capillaries (3-11 capillaries) were found scattered in the fibrosa and spongiosa with an average lumen area of 0.39 ± 0.18 mm(2). CONCLUSIONS: The blood vessels in atrioventricular valves also can be seen in the absence of inflammation and are likely to be a necessary component of valve leaflets. Thus, when performing procedures involving in situ tissue engineering and valve repair the physician needs to be aware of the presence of these vessels in human heart valves.

Morphometry of myenteric neurons in stomach.

The wall of the gastrointestinal tract presents extensive plexuses of nerve fibres and neuronal cell bodies responsible for the modulation of the rhythmic gastrointestinal peristaltic activities, among other functions. One of the most developed ganglionated plexuses of the gastrointestinal tract is the Myenteric plexus located between the inner circular layer and outer longitudinal layer of the smooth muscle tunica. The musculature of fundus, body and pyloric parts of stomach are differently disposed and they perform different functions. Thus the present study was conducted to study the myenteric plexus of all parts of stomach by counting the number of collections of neurons, number of neurons in each collection, diameter and area of the neurons of the plexus. The stomach walls of 1 cm in size were taken from 5 cadavers of medical post mortem cases from Postgraduate Institute of Medical Sciences and Research, Chandigarh and were processed for paraffin sections. 5 and 10 micro thick sections were stained with haematoxylin and Eosin and examined under light microscope. Randomly selected sections were photomicrographed using digital camera and morphometrical analysis was done using Image-Pro Express software. Number of collections of neurons was maximum in fundus with an average of 4.521 and each collection on an average contain 5.27 neurons ranging from 1-31, while body had 3.292 collections containing 1-19 neurons (mean: 3.198), pylorus had 3.883 collections of neurons which contained 1-16 neurons (mean: 4.411). The neurons were classified as small, medium and large according to the size of the area of their cell bodies. In this way, 11.3% neurons were found to be small, 69.5% medium and 19.1% large in fundus, 8.7% small, 80.6% medium and 11.2% large in body and 11.1% small, 74.3% medium and 14.5% large in pylorus.