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Mucosal vaccines protect against respiratory virus infection by stimulating the production of IgA antibodies that protect against virus invasion of the mucosal epithelium. In this study, a novel protein subunit mucosal vaccine was constructed for protection against infection by the beta coronavirus SARS-CoV-2. The vaccine was assembled by linking a gene encoding the SARS-CoV-2 virus S1 angiotensin converting enzyme receptor binding domain (ACE-2-RBD) downstream from a DNA fragment encoding the cholera toxin B subunit (CTB), a mucosal adjuvant known to stimulate vaccine immunogenicity. A 42 kDa vaccine fusion protein was identified in homogenates of transformed E. coli BL-21 cells by acrylamide gel electrophoresis and by immunoblotting against anti-CTB and anti-ACE-2-RBD primary antibodies. The chimeric CTB-SARS-CoV-2-ACE-2-RBD vaccine fusion protein was partially purified from clarified bacterial homogenates by nickel affinity column chromatography. Further vaccine purification was accomplished by polyacrylamide gel electrophoresis and electro-elution of the 42 kDa chimeric vaccine protein. Vaccine protection against SARS-CoV-2 infection was assessed by oral, nasal, and parenteral immunization of BALB/c mice with the CTB-SARS-CoV-2-ACE-2-RBD protein. Vaccine-induced SARS-CoV-2 specific antibodies were quantified in immunized mouse serum by ELISA analysis. Serum from immunized mice contained IgG and IgA antibodies that neutralized SARS-CoV-2 infection in Vero E6 cell cultures. In contrast to unimmunized mice, cytological examination of cell necrosis in lung tissues excised from immunized mice revealed no detectable cellular abnormalities. Mouse behavior following vaccine immunization remained normal throughout the duration of the experiments. Together, our data show that a CTB-adjuvant-stimulated CTB-SARS-CoV-2-ACE-2-RBD chimeric mucosal vaccine protein synthesized in bacteria can produce durable and persistent IgA antibodies in mice that neutralize the SARS-CoV-2 subvariant Omicron BA.1.1.

A chimeric protein vaccine composed of the cholera toxin B subunit fused to proinsulin (CTB-INS) was shown to suppress type 1 diabetes onset in NOD mice and upregulate biosynthesis of the tryptophan catabolic enzyme indoleamine 2, 3-dioxygenase (IDO1) in human dendritic cells (DCs). Here we demonstrate siRNA inhibition of the NF-κB-inducing kinase (NIK) suppresses vaccine-induced IDO1 biosynthesis as well as IKKα phosphorylation. Chromatin immunoprecipitation (ChIP) analysis of CTB-INS inoculated DCs showed that RelB bound to NF-κB consensus sequences in the IDO1 promoter, suggesting vaccine stimulation of the non-canonical NF-κB pathway activates IDO1 expression in vivo. The addition of Tumor Necrosis Factor Associated Factors (TRAF) TRAF 2, 3 and TRAF6 blocking peptides to vaccine inoculated DCs was shown to inhibit IDO1 biosynthesis. This experimental outcome suggests vaccine activation of the TNFR super-family receptor pathway leads to upregulation of IDO1 biosynthesis in CTB-INS inoculated dendritic cells. Together, our experimental data suggest the CTB-INS vaccine uses a TNFR-dependent signaling pathway of the non-canonical NF-κB signaling pathway resulting in suppression of dendritic cell mediated type 1 diabetes autoimmunity.

Purpose: The Veteran's Health Administration (VHA) has created a training program for interdisciplinary teams of providers on the unique treatment needs of transgender veterans. An overview of this program's structure and content is described along with an evaluation of each session and the program overall. Methods: A specialty care team delivered 14 didactic courses supplemented with case consultation twice per month over the course of 7 months through video teleconferencing to 16 teams of learners. Each team, consisting of at least one mental health provider (e.g., social worker, psychologist, or psychiatrist) and one medical provider (e.g., physician, nurse, physician assistant, advanced practice nurse, or pharmacist), received training and consultation on transgender veteran care. Results: In the first three waves of learners, 111 providers across a variety of disciplines attended the sessions and received training. Didactic topics included hormone therapy initiation and adjustments, primary care issues, advocacy within the system, and psychotherapy issues. Responses were provided to 39 veteran-specific consult questions to augment learning. Learners reported an increase in knowledge plus an increase in team cohesion and functioning. As a result, learners anticipated treating more transgender veterans in the future. Conclusion: VHA providers are learning about the unique healthcare needs of transgender veterans and benefitting from the training opportunity offered through the Transgender Specialty Care Access Network-Extension of Community Healthcare Outcomes program. The success of this program in training interdisciplinary teams of providers suggests that it might serve as a model for other large healthcare systems. In addition, it provides a path forward for individual learners (both within VHA and in the community) who wish to increase their knowledge., Competing Interests: No competing financial interests exist.

Rationale. To evaluate the effectiveness of group medical appointments (GMAs) for patients with type 2 diabetes. Objective. To compare A1C levels of patients participating in GMAs to those of patients who received usual primary care. Design and methods. This study was a retrospective electronic chart review comparing GMA care for 52 male patients to usual primary care for 52 male patients. Demographic (age, marital status, and ethnicity/race) and health-related (height, weight, BMI, duration of diabetes, use of alcohol and tobacco, and A1C) variables were analyzed. Results. A greater proportion of GMA patients (50%) versus usual primary care patients (19.2%) reached target A1C goals (P = 0.001). GMA participants also had a significantly faster rate of decline in A1C over time compared to usual primary care patients (P < 0.001). Conclusion. This study demonstrated that the concept of medical management delivered in a group approach had a positive effect on glycemic control in patients with type 2 diabetes. GMAs were found to be an effective approach to achieving patient-centered goals for improving the glycemic control of patients with type 2 diabetes.

Indoleamine 2, 3-dioxygenase (IDO) is the first and rate limiting catabolic enzyme in the degradation pathway of the essential amino acid tryptophan. By cleaving the aromatic indole ring of tryptophan, IDO initiates the production of a variety of tryptophan degradation products called "kynurenines" that are known to exert important immuno-regulatory functions. Because tryptophan must be supplied in the diet, regulation of tryptophan catabolism may exert profound effects by activating or inhibiting metabolism and immune responses. Important for survival, the regulation of IDO biosynthesis and its activity in cells of the immune system can critically alter their responses to immunological insults, such as infection, autoimmunity and cancer. In this review, we assess how IDO-mediated catabolism of tryptophan can modulate the immune system to arrest inflammation, suppress immunity to cancer and inhibit allergy, autoimmunity and the rejection of transplanted tissues. Finally, we examine how vaccines may enhance immune suppression of autoimmunity through the upregulation of IDO biosynthesis in human dendritic cells.

Dendritic cells (DC) interact with naïve T cells to regulate the delicate balance between immunity and tolerance required to maintain immunological homeostasis. In this study, immature human dendritic cells (iDC) were inoculated with a chimeric fusion protein vaccine containing the pancreatic β-cell auto-antigen proinsulin linked to a mucosal adjuvant the cholera toxin B subunit (CTB-INS). Proteomic analysis of vaccine inoculated DCs revealed strong up-regulation of the tryptophan catabolic enzyme indoleamine 2, 3-dioxygenase (IDO1). Increased biosynthesis of the immunosuppressive enzyme was detected in DCs inoculated with the CTB-INS fusion protein but not in DCs inoculated with proinsulin, CTB, or an unlinked combination of the two proteins. Immunoblot and PCR analyses of vaccine treated DCs detected IDO1mRNA by 3 hours and IDO1 protein synthesis by 6 hours after vaccine inoculation. Determination of IDO1 activity in vaccinated DCs by measurement of tryptophan degradation products (kynurenines) showed increased tryptophan cleavage into N-formyl kynurenine. Vaccination did not interfere with monocytes differentiation into DC, suggesting the vaccine can function safely in the human immune system. Treatment of vaccinated DCs with pharmacological NF-κB inhibitors ACHP or DHMEQ significantly inhibited IDO1 biosynthesis, suggesting a role for NF-κB signaling in vaccine up-regulation of dendritic cell IDO1. Heat map analysis of the proteomic data revealed an overall down-regulation of vaccinated DC functions, suggesting vaccine suppression of DC maturation. Together, our experimental data indicate that CTB-INS vaccine induction of IDO1 biosynthesis in human DCs may result in the inhibition of DC maturation generating a durable state of immunological tolerance. Understanding how CTB-INS modulates IDO1 activity in human DCs will facilitate vaccine efficacy and safety, moving this immunosuppressive strategy closer to clinical applications for prevention of type 1 diabetes autoimmunity.

High levels of serum long chain saturated fatty acids (LCSFAs) have been associated with inflammation in type 2 diabetes. Dietary SFAs can promote inflammation, the secretion of IgG antibodies, and secretion of the proinflammatory cytokine IL-1β. This study characterizes anti-LCSFA IgG antibodies from patients with type 2 diabetes. Serum samples from several cohorts with type 2 diabetes were analyzed for the presence of anti-LCSFA IgG, the cytokine IL-1β, and nonesterified fatty acids. Anti-LCSFA IgG was isolated from patient samples and used for in vitro characterization of avidity and specificity. A cohort participating in En Balance, a diabetes health education program that improved diabetes management, tested positive for anti-LCSFA IgG. Following the 3-month program, the cohort showed a significant reduction in anti-LCSFA IgG levels. Anti-LCSFA antibodies isolated from these patients demonstrated high avidity, were specific for long chain SFAs, and correlated with serum fatty acids in patients with managed type 2 diabetes. Interestingly, anti-LCSFA IgG neutralized PA-induced IL-1β secretion by dendritic cells. Our data shows that nonesterified SFAs are recognized by IgG antibodies present in human blood. The identification of anti-LCSFA IgG antibodies in human sera establishes a basis for further exploration of lipid induced immune responses in diabetic patients.

Adipose tissue secretes the adipokine, adiponectin (ADPN), which increases insulin sensitivity. Because some of the metabolic effects of exercise and ADPN are similar, exercise has been proposed to increase ADPN. However, most short-term (≤3 mos) and constant-effort exercise protocols have not produced increases in ADPN. Furthermore, no direct comparisons of male and female subjects on the effect of exercise on ADPN levels have been reported. We hypothesized that long-term (6 mos), progressive training would increase ADPN levels in both males and females. We recruited middle-aged, untrained males and females to participate in an interventional study employing a marathon training regimen progressing from 9.7 to 88.5 km (6 to 55 miles) per week over 6 mos. At baseline, we matched the mean ages of the male and female groups. We collected and stored fasting plasma samples and recorded body measurements at 0 (baseline) and 6 mos. Stored samples were analysed for insulin, glucose, and ADPN. ADPN increased significantly among both males (from 5.89 ± 2.46 (mean ± SD) to 7.65 ± 3.18 μg/ml; p < 0.05) and females (from 8.48 ± 3.22 to 10.56 ± 4.05 μg/ml; p < 0.05). The extent of the increase in ADPN was similar in the male (40.7 ± 50%; median, 12.1%) and female (27.0 ± 31.1%; median, 22.3%) groups. However, there was no significant reduction in insulin resistance as measured by the HOMA-IR scores in either group. We conclude that long-term, progressive aerobic training increases circulating ADPN levels in middle-aged, untrained males and females.

PTH clearly plays a role in maintaining the hypercalcemia of familial benign hypercalcemia (FBH or familial hypocalciuric hypocalcemia). To better define the abnormalities of parathyroid function in FBH and primary hyperparathyroidism (1 degree HPT), we used a two-site immunochemiluminometric assay for intact PTH to examine PTH suppressibility in normal individuals and patients having FBH or 1 degree HPT. Twelve normal, 11 FBH, and 7 1 degree HPT subjects were given calcium (Ca) iv with frequent sampling for ionized Ca and intact PTH. In normal and FBH subjects, plasma PTH levels decreased essentially identically in response to iv Ca. In the 1 degree HPT group, PTH was not normally suppressible. However, there was a spectrum of responsiveness in 1 degree HPT patients, with a significant correlation between tumor mass and degree of PTH nonsuppressibility (r = 0.87, P = 0.01). Analysis of the relationship between plasma PTH and ionized Ca values in the three groups demonstrated a shift to the right in the FBH curve, with no difference of slope, consistent with the notion of a simple "set-point" error in FBH. In contrast, the curve in 1 degree HPT was not only shifted to the right but also differed from normal in slope (normal, -8.92; 1 degree HPT, -3.92, P = 0.04). Thus, we propose that the parathyroid functional abnormality in FBH represents a simple set-point error, whereas the defect in 1 degree HPT consists of a set-point error combined with varying degrees of Ca nonsuppressible PTH secretion that may be related to tumor mass.

Humoral hypercalcemia of malignancy (HHM) is at least partly caused by tumor secretion of PTH-related peptide (PTHrP), but there is growing evidence for cosecretion with PTHrP of other bone-resorbing peptides, such as the cytokine interleukin-1 alpha (IL-1 alpha). Administration of PTHrP in vivo and in vitro generally mimics the actions of PTH itself, with increases in both resorption and formation of bone. However, bone in HHM is characterized by uncoupling of bone turnover, with increased resorption and decreased formation. We performed experiments to determine whether IL-1 alpha might alter the effects of PTHrP and produce uncoupling. Thus, we administered to 100-g male rats by sc osmotic minipumps synthetic PTHrP-(1-34) alone (2 micrograms/100 g/day), recombinant IL-1 alpha alone (1.5 micrograms/100 g/day), both peptides together at the previous doses, or vehicle only. We infused 5 groups of 12 rats each (PTHrP, IL-1 alpha, PTHrP plus IL-1 alpha, ad libitum fed control, and controls pair-fed to the PTHrP plus IL-1 alpha group) for 14 days. At the end of the study, blood and urine were taken for chemical measurements, and tibias and femurs were harvested for histomorphometry and extraction of RNA from periosteal cells. As expected, PTHrP induced hypercalcemia, relative hypophosphatemia, phosphaturia, and reduced bone mass. Osteoblast number was increased, but osteoclast number was not. Indices of bone formation were unchanged or reduced. The dose of IL-1 alpha chosen had no statistically significant effect, except for reduced longitudinal bone growth, but when combined with PTHrP, IL-1 alpha reduced hypercalcemia, hypophosphatemia, and phosphaturia. In contrast to the blood and urine effects, IL-1 alpha did not interact significantly with PTHrP's effect on bone measurements. Northern analysis of periosteal cell mRNA showed that PTHrP reduced expression of osteocalcin, but not glyceraldehyde-3-phosphate dehydrogenase; IL-1 alpha had no additional effect. These data suggest that 1) continuously administered PTHrP alone may induce uncoupled bone turnover with decreased cortical bone formation; 2) IL-1 alpha appears to inhibit strongly the renal effects of PTHrP and weakly (if at all) its actions on bone and, thus, to decrease its hypercalcemic, phosphaturic, and hypophosphatemic actions; and 3) cosecretion of IL-1 alpha, and possibly other peptide cytokines, with PTHrP may modify the clinical expression of HHM.

Plasma concentrations of PTH are much lower for a given calcium or phosphorus level in patients with familial benign hypercalcemia (FBH, or familial hypocalciuric hypercalcemia) than in those with primary hyperparathyroidism; these and other data suggest that there might be tissue hypersensitivity to PTH in FBH. To test this hypothesis, we have used cultured dermal fibroblasts from abdominal skin biopsies of six patients with FBH and six age- and sex-matched controls as surrogate PTH-responsive tissues. Cells in 24-well plastic plates were exposed to vehicle, human PTH-(1-34) (10(-10)-10(-7) M), prostaglandin E2 (10(-6) M), or isoproterenol (10(-4) M) for 10 min in the presence of isobutylmethylxanthine, and cellular cAMP was determined by RIA. All cells responded to PTH with dose-dependent increases in cAMP, and all responded strongly to prostaglandin E2 and isoproterenol. There were no consistent or significant differences between control and FBH fibroblasts in maximal responses to the three agonists, and half-maximal stimulation was achieved with about 10(-9) M PTH in both normal and FBH cells. These data are not consistent with increased tissue sensitivity to PTH in FBH.

The cause of hypercalcemia in familial benign hypercalcemia (FBH; also called familial hypocalciuric hypercalcemia) is unclear, although it is PTH dependent. It is also uncertain how plasma PTH levels are related to the severity of biochemical abnormalities in FBH. Because the PTH-related peptide (PTHrP) has many PTH-like actions, it might have a role in the hypercalcemia of FBH. Thus, we studied 29 patients with FBH from 11 families, 29 age- and sex-matched controls, and 42 patients with primary hyperparathyroidism (1 degree HPT), measuring PTH with a highly sensitive two-site immunochemiluminometric assay and the hypercalcemic tumor factor PTH-related peptide (PTHrP) with an extraction/concentration RIA. Plasma PTH values were elevated in 86% of 1 degree HPT patients (36 of 42), but in only 20% of FBH patients, (6 of 29). Plasma PTHrP was elevated in 1 FBH patient, and the group mean value was normal. Plasma PTH was positively correlated with calcium (Ca) in 1 degree HPT (r = 0.66; P less than 0.0001) and in FBH (r = 0.53; P less than 0.004), but the slopes of the regressions were markedly different: 1 degree HPT, 6.72; FBH, 1.61 (P less than 0.0001). There was a negative correlation between PTH and phosphorus (P) in 1 degree HPT (r = -0.39; P less than 0.01) and in FBH (r = -0.41; P less than 0.03), but, again, the slopes differed greatly: 1 degree HPT, -6.57; FBH, -1.95 (P less than 0.0001). There were no correlations between PTHrP and Ca or between PTH and PTHrP. The sums and products of PTH and PTHrP were not better correlated with Ca than PTH alone. Thus, PTH values are lower at given Ca and P levels in patients with FBH than in those with 1 degree HPT, suggesting that PTH is more effective in raising Ca and lowering P in FBH than in 1 degree HPT. The enigma of FBH remains: what molecular defect can simultaneously cause parathyroid cell insensitivity to Ca, enhanced renal tubular reabsorption of Ca, increased renal rejection of P, and enhanced or retained sensitivity to PTH?

We have investigated the effects of PTH-induced desensitization on second messenger interactions in the rat osteosarcoma cell line ROS 17/2.8. Adenylate cyclase activation was assessed by accumulation of immunoassayable cAMP, and cytosolic calcium ion ([Ca2+]i) concentrations were measured in adherent perifused cells loaded with the Ca2(+)-sensitive bioluminescent protein aequorin. Preexposure to rat PTH-(1-34) [rPTH-(1-34); 10(-8) M for 48 h, then 10(-7) M for 24 h] dramatically reduced (by 85%) the cAMP response to fresh challenge [2 min; 10(-9)-10(-7) M rPTH-(1-34)], but the peak PTH-induced rise of [Ca2+]i was not diminished significantly (0-20%). Nevertheless, we did observe other changes in the PTH-induced [Ca2+]i response. Exposure of treated cells to (Bu)2cAMP nearly abolished the [Ca2+]i response to PTH (greater than 80% reduction), but had much less effect on the PTH-stimulated [Ca2+]i increment of the naive cells (less than 35% reduction). Treated cells also had a blunted [Ca2+]i response to PTH in the presence of low extracellular calcium (greater than 60% reduction), but in the naive cells, low extracellular Ca2+ did not significantly diminish the peak PTH-induced [Ca2+]i rise, although low extracellular Ca2+ dramatically reduced the area under this [Ca2+]i transient (greater than 50%). Low extracellular Ca2+ had no influence on the peak [Ca2+]i responses of treated cells to bradykinin or prostaglandin F2 alpha. Although the peak PTH-stimulated [Ca2+]i rise of treated cells in normal Ca2+ medium was not significantly attenuated, the time to half-maximum [Ca2+]i concentration was significantly increased (greater than 100%), and the area under the [Ca2+]i transient was diminished. These alterations in the [Ca2+]i response of treated cells were not observed upon challenge with bradykinin or prostaglandin F2 alpha. Thus, 1) the cAMP and [Ca2+]i responses of ROS 17/2.8 cells to rPTH-(1-34) are not obligatorily coupled; 2) the response of naive cells to PTH includes both the release of Ca2+ from intracellular stores and the entry of extracellular Ca2+; and 3) pretreatment of these cells with rPTH-(1-34) augments the dependence on Ca2+ entry during hormone rechallenge. We propose that the preserved PTH-stimulated [Ca2+]i rise in treated cells results partly from loss of cAMP-mediated inhibition of extracellular Ca2+ entry.

Specific insulin-like growth factor-binding proteins (IGFBPs) that may enhance or inhibit insulin-like growth factor (IGF) action are produced in various tissues. In the present study we demonstrated that IGFBPs are synthesized and secreted by rat osteoblast-like cells (UMR 106-01). PTH and PTH-related peptide (PTHrP) were potent stimuli for IGFBP production by UMR cells, whereas GH, IGF-I, insulin, epidermal growth factor, and T3 had little or no effect. A maximal 8- to 30-fold increase in IGFBP production was attained at 10(-7)-10(-6) M PTH and PTHrP, with a half-maximal effect at approximately 10(-9) M. By Western blot analysis, PTH and PTHrP markedly and selectively increased the production of 29,000 mol wt (Mr) and, to a lesser extent, 24,000 Mr IGFBPs. Agents that elevate intracellular cAMP by different mechanisms [(Bu)2cAMP, forskolin, and isobutylmethylxanthine] mimicked the effect of PTH and PTHrP on IGFBP synthesis. In comparison, PTH did not stimulate IGFBP production in fibroblasts and ROS 17/2.8 cells, which secrete IGFBPs of 42,000, 38,000, 34,000, 28,000, and 24,000 Mr, but not of 29,000 Mr. The PTH-responsive IGFBPs from UMR cells were nonglycosylated proteins with preferential affinity for IGF-I over IGF-II. These IGFBPs were not immunoprecipitated with antisera against rat IGFBP-2 or human IGFBP-1. Thus, PTH and PTHrP increase the production in UMR 106-01 cells of discrete IGFBP forms with Mr of 29,000 and 24,000 through a cAMP-mediated mechanism, independent of IGF-I synthesis. Taken with the known effects of PTH on IGF production in bone cells, the data suggest that PTH and PTHrP may modulate local IGF action in bone through the regulation of specific IGFBP availability.

Administration of calcitonin (CT) or parathyroid hormone (PTH) in rats, induced phosphaturia with a concomitant increase in urinary excretion of cyclic AMP. CT produced a rise in arterial cAMP and filtered cAMP, with no increase in urinary nephrogenous cAMP. PTH on the other hand, produced an elevation in urinary nephrogenous cAMP with no increase in arterial cAMP and filtered cAMP. These results indicate that the source of urinary cAMP after CT is non-renal, whereas after PTH it is renal in origin.

Alcohol ingestion during pregnancy is known to cause fetal malformation and growth retardation. We investigated the effect of alcohol on mineral content and fetal development in rats fed 24% (v/v) alcohol eight weeks prior to and during pregnancy. Rats ingesting alcohol produced fewer fetuses (6.3 +/- 0.3 vs 9.6 +/- 0.3 in control) with lower fetal weight (3.48 +/- 0.09 vs 4.12 +/- 0.08 gm in control) and heavier placentas (0.66 +/- 0.05 vs 0.50 +/- 0.01 gm in control). The fetuses of alcoholic rats contained lower zinc (423.8 +/- 4.5 vs 459.9 +/- 5.4 microEq/100 gm dry weight in control) and magnesium (12.4 +/- 0.1 vs 12.7 +/- 0.1 mEq/100 gm dry weight in control) in the total carcass.

The response to parathyroid hormone (PTH) and calcitonin (CT) was studied in eight children with various bone diseases by determining the serum calcium (Ca) and phosphate (P) concentration, urinary phosphate excretion rate, renal phosphate clearance, the percentage of filtered phosphate reabsorbed by the renal tubule (%TRP), creatinine clearance (Ccr), urinary cyclic adenosine 3',5' monophosphate excretion rate (UcGMPV). Administration of PTH caused no significant change in serum Ca and P values, whereas CT produced a decrease in Ca (delta Ca, -1.4 +/- 0.1 mg/100 ml) and P (delta P; -1.1 +/- 0.1 mg/100 ml). There was an increase in UcAMP V (delta UcAMP; 437 +/- 74 nmoles/min/100 ml Ccr) without any significant change in UcGMPV after administration of PTH. Phosphaturia was produced by both PTH (delta TRP, -18 +/- 3%) and CT (delta TRP, -13 +/- 2%). However, CT did not elicit any increase in either UcAMPV or UcGMPV.

The effect of hypomagnesemia on renal concentrating ability was assessed in rats fed diets either low in magnesium or low in magnesium and calcium for 30 days. The rats fed a low-magnesium diet became hypomagnesemic (0.26 +/- 0.03 versus 1.53 +/- 0.04 mEq/L in controls), hypercalcemic (5.96 +/- 0.04 versus 5.22 +/- 0.11 mEq/L in controls), and hypokalemic (3.1 +/- 0.1 versus 4.2 +/- 0.4 mEq/L in controls) with decreased muscle content of potassium. Despite being hypomagnesemic, hypercalcemic, and potassium depleted, the rats had normal renal concentration ability (2499 +/- 65 versus 2415 +/- 119 mOsm/kg H2O in control). Those rats fed a diet low in both magnesium and calcium became hypomagnesemic (0.41 +/- 0.08 versus 1.53 +/- 0.04 mEq/L in controls) but were hypocalcemic. They also had normal renal concentrating ability (2399 +/- 109 versus 2415 +/- 119 mOsm/kg H2O in controls). It is concluded that hypomagnesemia does not decrease renal concentrating ability in rats. Furthermore, a normal concentrating ability demonstrated in hypomagnesemic rats, in spite of hypercalcemia and potassium depletion suggests that hypomagnesemia may ameliorate the deleterious effects of hypercalcemia and/or potassium depletion on renal concentrating ability.