Vascular Inflammatory Cells in Hypertension

Hypertension is a common disorder with uncertain etiology. In the last several years, it has become evident that components of both the innate and adaptive immune system play an essential role in hypertension. Macrophages and T cells accumulate in the perivascular fat, the heart and the kidney of hypertensive patients, and in animals with experimental hypertension. Various immunosuppressive agents lower blood pressure and prevent end-organ damage. Mice lacking lymphocytes are protected against hypertension, and adoptive transfer of T cells, but not B cells in the animals restores their blood pressure response to stimuli such as angiotensin II or high salt. Recent studies have shown that mice lacking macrophages have blunted hypertension in response to angiotensin II and that genetic deletion of macrophages markedly reduces experimental hypertension. Dendritic cells have also been implicated in this disease. Many hypertensive stimuli have triggering effects on the central nervous system and signals arising from the circumventricular organ seem to promote inflammation. Studies have suggested that central signals activate macrophages and T cells, which home to the kidney and vasculature and release cytokines, including IL-6 and IL-17, which in turn cause renal and vascular dysfunction and lead to blood pressure elevation. These recent discoveries provide a new understanding of hypertension and provide novel therapeutic opportunities for treatment of this serious disease

The Impact of Hyperosmolality on Activation and Differentiation of B Lymphoid Cells

B lymphocytes, as a central part of adaptive immune responses, have the ability to fight against an almost unlimited numbers of pathogens. Impairment of B cell development, activation and differentiation to antibody secreting plasma cells can lead to malignancy, allergy, autoimmunity and immunodeficiency. However, the impact of environmental factors, such as hyperosmolality or osmotic stress caused by varying salt concentrations in different lymphoid organs, on these processes is not well-understood. Here, we report that B cells respond to osmotic stress in a biphasic manner. Initially, increased osmolality boosted B cell activation and differentiation as shown by an untimely downregulation of Pax5 as well as upregulation of CD138. However, in the second phase, we observed an increase in cell death and impaired plasmablast differentiation. Osmotic stress resulted in impaired class switch to IgG1, inhibition of phosphorylation of p38 mitogen-activated kinase and a delayed NFAT5 response. Overall, these findings demonstrate the importance of microenvironmental hyperosmolality and osmotic stress caused by NaCl for B cell activation and differentiation

High-salt diet causes osmotic gradients and hyperosmolality in skin without affecting interstitial fluid and lymph

The common notion is that the body Na+ is maintained within narrow limits for fluid and blood pressure homeostasis. Several studies have, however, shown that considerable amounts of Na+ can be retained or removed from the body without commensurate water loss and that the skin can serve as a major salt reservoir. Our own data from rats have suggested that the skin is hypertonic compared with plasma on salt storage and that this also applies to skin interstitial fluid. Even small electrolyte gradients between plasma and interstitial fluid would represent strong edema-generating forces. Because the water accumulation has been shown to be modest, we decided to reexamine with alternative methods in rats whether interstitial fluid is hypertonic during salt accumulation induced by high-salt diet (8% NaCl and 1% saline to drink) or deoxycorticosterone pellet implantation. These treatments resulted both in increased systemic blood pressure, skin salt, and water accumulation and in skin hyperosmolality. Interstitial fluid isolated from implanted wicks and lymph draining the skin was, however, isosmotic, and Na+ concentration in fluid isolated by centrifugation and in lymph was not different from plasma. Interestingly, by eluting layers of the skin, we could show that there was an osmolality and urea gradient from epidermis to dermis. Collectively, our data suggest that fluid leaving the skin as lymph is isosmotic to plasma but also that the skin can differentially control its own electrolyte microenvironment by creating local gradients that may be functionally important.acceptedVersio

Ex Vivo High Salt Activated Tumor-Primed CD4+T Lymphocytes Exert a Potent Anti-Cancer Response

Cell based immunotherapy is rapidly emerging as a promising cancer treatment. A modest increase in salt (sodium chloride) concentration in immune cell cultures is known to induce inflammatory phenotypic differentiation. In our current study, we analyzed the ability of salt treatment to induce ex vivo expansion of tumor-primed CD4 (cluster of differentiation 4)+T cells to an effector phenotype. CD4+T cells were isolated using immunomagnetic beads from draining lymph nodes and spleens from tumor bearing C57Bl/6 mice, 28 days post-injection of Py230 syngeneic breast cancer cells. CD4+T cells from non-tumor bearing mice were isolated from splenocytes of 12-week-old C57Bl/6 mice. These CD4+T cells were expanded ex vivo with five stimulation cycles, and each cycle comprised of treatment with high salt (Δ0.035 M NaCl) or equimolar mannitol controls along with anti-CD3/CD28 monoclonal antibodies for the first 3 days, followed by the addition of interleukin (IL)-2/IL-7 cytokines and heat killed Py230 for 4 days. Ex vivo high salt treatment induced a two-fold higher Th1 (T helper type 1) expansion and four-fold higher Th17 expansion compared to equimolar mannitol treatment. Importantly, the high salt expanded CD4+T cells retained tumor-specificity, as demonstrated by higher in vitro cytotoxicity against Py230 breast cancer cells and reduced in vivo syngeneic tumor growth. Metabolic studies revealed that high salt treatment enhanced the glycolytic reserve and basal mitochondrial oxidation of CD4+T cells, suggesting a role of high salt in enhanced pro-growth anabolic metabolism needed for inflammatory differentiation. Mechanistic studies demonstrated that the high salt induced switch to the effector phenotype was mediated by tonicity-dependent transcription factor, TonEBP/NFAT5. Using a transgenic murine model, we demonstrated that CD4 specific TonEBP/NFAT5 knock out (CD4cre/creNFAT5flox/flox) abrogated the induction of the effector phenotype and anti-tumor efficiency of CD4+T cells following high salt treatment. Taken together, our data suggest that high salt-mediated ex vivo expansion of tumor-primed CD4+T cells could induce effective tumor specific anti-cancer responses, which may have a novel cell-based cancer immunotherapeutic application

High salt induces P-glycoprotein mediated treatment resistance in breast cancer cells through store operated calcium influx

Recent evidence from our laboratory has demonstrated that high salt (Δ0.05 M NaCl) induced inflammatory response and cancer cell proliferation through salt inducible kinase-3 (SIK3) upregulation. As calcium influx is known to effect inflammatory response and drug resistance, we examined the impact of high salt on calcium influx in breast cancer cells. Treatment of MCF-7 and MDA-MB-231 cells with high salt induced an enhanced intracellular calcium intensity, which was significantly decreased by store operated calcium entry (SOCE) inhibitor co-treatment. Further, high salt induced P-glycoprotein (P-gp) mediated paclitaxel drug resistance in breast cancer cells. Murine tumor studies demonstrated that injection of MCF-7 cells cultured in high salt, exerted higher tumorigenicity compared to the basal cultured counterpart. Knock down of SIK3 by specific shRNA inhibited tumorigenicty, expression of SOCE regulators and P-gp activity, suggesting SIK3 is an upstream mediator of SOCE induced calcium influx. Furthermore, small molecule inhibitor, prostratin, exerted anti-tumor effect in murine models through SIK3 inhibition. Taken together, we conclude that SIK3 is an upstream regulator of store operated calcium entry proteins, Orai1 and STIM1, and mediates high salt induced inflammatory cytokine responses and P-gp mediated drug resistance. Therefore, small molecule inhibitors, such as prostratin, could offer novel anti-cancer approaches

Dietary Salt Accelerates Orthodontic Tooth Movement by Increased Osteoclast Activity

Dietary salt uptake and inflammation promote sodium accumulation in tissues, thereby modulating cells like macrophages and fibroblasts. Previous studies showed salt effects on periodontal ligament fibroblasts and on bone metabolism by expression of nuclear factor of activated T-cells-5 (NFAT-5). Here, we investigated the impact of salt and NFAT-5 on osteoclast activity and orthodontic tooth movement (OTM). After treatment of osteoclasts without (NS) or with additional salt (HS), we analyzed gene expression and the release of tartrate-resistant acid phosphatase and calcium phosphate resorption. We kept wild-type mice and mice lacking NFAT-5 in myeloid cells either on a low, normal or high salt diet and inserted an elastic band between the first and second molar to induce OTM. We analyzed the expression of genes involved in bone metabolism, periodontal bone loss, OTM and bone density. Osteoclast activity was increased upon HS treatment. HS promoted periodontal bone loss and OTM and was associated with reduced bone density. Deletion of NFAT-5 led to increased osteoclast activity with NS, whereas we detected impaired OTM in mice. Dietary salt uptake seems to accelerate OTM and induce periodontal bone loss due to reduced bone density, which may be attributed to enhanced osteoclast activity. NFAT-5 influences this reaction to HS, as we detected impaired OTM and osteoclast activity upon deletion