Calcium oxalate crystal formation in patients with hyperparathyroidism and hyperthyroidism and related metabolic disturbances

The crystallization of calcium oxalate in the urine of patients with hyperparathyroidism and hyperthyroidism was studied using a mixed suspension mixed product removal (MSMPR) system. In addition, calcium metabolism in hyperthyroidism and its relationship to urolithiasis was investigated. The urines from all the three groups (normal subjects, hyperparathyroid and hyperthyroid patients) showed reduced nucleation rates and increased growth rates in comparison with the control synthetic urine. The nucleation rate was not significantly different between the three human urine groups, while the growth rate was significantly higher in the hyperparathyroid group compared to the normal and hyperthyroid groups. Crystal volume (suspension density) in the hypetparathyroid group was approximately twice that in the other two groups. Serum and ionized calcium levels in hyperparathyroid patients were higher than in normal subjects, while hyperthyroid patients had levels only slightly higher than those in normal subjects. The hyperparathyroid and hyperthyroid groups differed significantly from the normal group in urinary calcium excretion. These two groups also showed significantly higher levels of serum alkaline phosphatase and urinary hydroxyproline than did the normal group. Although hyperthyroid patients have a calcium metabolism similar to hyperparathyroid patients, the incidence of urolithiasis is no different between hyperthyroid and normal subjects. The results of both crystallization and calcium metabolism in hyperparathyroid patients were not significantly different between those with and without urolithiasis. The result of crystallization was also not significantly different between hyperparathyroid patients with and without hypercalciuria. This study suggests that hypercalciuria alone does not produce urinary stones and that urine from hyperparathyroid patients may contain promotors of calcium oxalate crystallization and calcium stone formation.     

Changes in aldosterone and cortisol secretion and serum thyroxine levels in patients with active urolithiasis

The changes in calcemia and calciuria levels following low calcium diet have been studied in 35 patients with active urolithiasis and in 20 healthy subjects. Blood serum concentrations of thyroxine, cortisol and aldosterone in basal conditions as well as cortisol and aldosterone following stimulation with synacten were determined in addition. The levels of calcemia and calciuria (2.56 +/- 0.015 mmol/l and 4.70 +/- 0.41 mmol/10 mmoles of creatinine, respectively) were found to be significantly higher in patients with active urolithiasis than in healthy subjects. In addition, in patients with urolithiasis the basal blood serum concentrations of thyroxine and aldosterone were significantly higher than in healthy subjects, while the reactivity of cortisol and aldosterone secretion to synacten stimulation was normal. The results obtained suggest the participation of the described hormonal aberrations in the pathogenesis of active urolithiasis.     

Protein abnormalities in adult respiratory distress syndrome, tuberculosis, and cystic fibrosis sera

Crossed immunoelectrophoresis (X-IEP) revealed several abnormalities in serum proteins from patients with adult respiratory distress syndrome (ARDS), tuberculosis (TB), and cystic fibrosis (CF). The two quite different kinds of pulmonary disease, one acute (ARDS) and the other chronic (TB and CF) exhibited serum changes specific for each disease and abnormalities associated with inflammation and pathogenesis, in general. In ARDS sera, most proteins were extremely low, presumably due to leakage into the lungs through damaged tissue, while the acute-phase proteins, orosomucoid, alpha 1-antitrypsin, alpha 1-antichymotrypsin, and haptoglobin, were markedly high when compared to the overall protein pattern. The extremely high alpha 1-antichymotrypsin values were not seen in corresponding TB and CF sera. Numerous TB patients had elevated alpha 1-antitrypsin, alpha 1-antichymotrypsin, and haptoglobin, but only the alpha 1-antitrypsin population mean was significantly different from normal. Gc-globulin, ceruloplasmin, and beta-lipoprotein were higher and alpha 1-lipoprotein and inter-alpha-trypsin inhibitor lower than normal. All other quantitative serum changes were not statistically significant. Surprisingly, all TB patients belonged to the Gc-1-1 genotype in contrast to the Gc-1-1, Gc-1-2, Gc-2-2 polymorphisms of the other populations. CF homozygote sera revealed statistically significant increases in the acute-phase proteins, alpha 1-antitrypsin, alpha 1-antichymotrypsin, and haptoglobin, while orosomucoid, transferrin, IgA, and IgG tended to be higher than normal. The tendency for higher levels of transferrin indicated possible iron deficiency in some patients. In contrast, prealbumin, alpha 1-lipoprotein, and inter-alpha-trypsin inhibitor were significantly depressed in CF patients. CF heterozygotes shared the decrease of alpha 1-lipoprotein with the patients while exhibiting small but significant depressions of alpha 2-macroglobulin and IgG. Though not statistically significant, lowered concentrations of alpha 1-antitrypsin were evident for the heterozygotes.     

Comparison of calcium oxalate values as indicators of risk for calcium oxalate calcinosis and the degree of urine saturation in patients with calcium urolithiasis and in healthy patients

Daily excretion of calcium, magnesium, oxalates, and citrates together with daily urine output were determined in 37 patients with calcium urolithiasis and in 25 healthy individuals. Basing on the obtained values, a degree of urine saturation with calcium oxalate with Marshall and Robertson technique and a value of risk factor with Tiselius technique were calculated. It was found that daily diuresis and excretion of calcium with the urine are significantly higher in patients with urolithiasis where as daily excretion of citrates with the urine is significantly lower than in healthy individuals. Risk index proved two-fold higher in the examined patients than in the healthy individuals (p greater than 0.001) while the degree of urine saturation with calcium oxalate did not differ significantly in both groups. The authors, discussing causes of seemingly different changes in both tested parameters, stressed diagnostic value of risk index which includes excretion of crystallization inhibitors (magnesium, citrates) and contrary to the degree of urine saturation is independent of daily urine output.     

The Distribution of Several Elements in Cat Urine and the Relation Between the Content of Elements and Urolithiasis

The concentrations of elements in urine obtained from cats with urolithiasis were compared with those of healthy cats. The concentration of several elements, such as sodium (Na), phosphorus (P), sulfur (S), and potassium (K), in urine obtained from cats with urolithiasis was significantly higher than that of healthy cats. A significant correlation (p<0.01) was found between the concentration of magnesium (Mg) and that of other elements, such as P (r=0.8913), S (r=0.6817), and K (r=0.8391), in the urine obtained from healthy cats. A significant correlation (r=0.7422, p<0.05) was also obtained between the concentration of K and that of P in urine collected from cats with urolithiasis, but the slope of regression line was significantly different from that of the urine obtained from healthy cats. Other correlations observed in healthy cats were not obtained from cats with urolithiasis. However, a significant correlation between the concentration of magnesium (Mg) and that of calcium was obtained only from cats with urolithiasis. The results of the present study suggest that urinary concentrations of various elements in cats with urolithiasis are higher than those of healthy cats. Furthermore, the balance of elements in the urine of cats with urolithiasis was altered.     

Urothelial Dysfunction and Increased Suburothelial Inflammation of Urinary Bladder Are Involved in Patients with Upper Urinary Tract Urolithiasis – Clinical and Immunohistochemistry Study

Objectives

To investigate the urothelial dysfunction and inflammation of urinary bladder in patients with upper urinary tract (UUT) urolithiasis through the results of cystoscopic hydrodistension and immunohistochemistry study.

Methods

Ninety-one patients with UUT urolithiasis underwent cystoscopic hydrodistension before the stone surgery. Immunofluorescence staining of E-cadherin, zonula occludens-1 (ZO-1), tryptase (mast cell activation), and TUNEL (urothelial apoptosis) were performed in 42 patients with glomerulations after hydrodistension, 10 without glomerulations, and 10 controls.

Results

Of the 91 patients, 62 (68.2%) developed glomerulations after hydrodistension. Lower urinary tract symptoms (LUTS) were present in 53.8% patients, in whom significantly smaller maximal anesthetic bladder capacity (MBC) was noted. Patients with middle or lower 1/3 ureteral stones had a significantly higher glomerulation rate (88.6% vs. 55.4%, p<0.01) and lower MBC (618.4±167.6 vs. 701.2±158.4 ml, p = 0.027) than those with upper 1/3 ureteral or renal stones. Patients with UUT urolithiasis had significantly lower expression of E-cadherin (26.2±14.8 vs. 42.4±16.7) and ZO-1 (5.16±4.02 vs. 11.02±5.66); and higher suburothelial mast cell (13.3±6.8 vs. 1.3±1.2) and apoptotic cell (2.6±2.5 vs. 0.1±0.3) numbers than in controls (all p<0.01).

Conclusions

Urothelial dysfunction and increased suburothelial inflammation and apoptosis are highly prevalent in the bladders of UUT urolithiasis patients, indicating inflammation cross-talk between UUT and urinary bladder. Patients with UUT urolithiaisis concomitant with LUTS had a smaller MBC, which may explain the presence of irritative bladder symptoms.     

Macrophage inflammatory protein-1α: A link between innate immunity and familial mediterranean fever?

The aim of this study is to investigate the relationship between chemokines and the inflammation in Familial Mediterranean Fever (FMF). Forty-nine patients with FMF (41 in remission and 8 in acute attack period) and 20 healthy controls were included in the study. Serum levels of macrophage inflammatory protein-1alpha (MIP-1alpha) were assessed in the patients and the controls, along with other parameters of disease activity, i.e., fibrinogen, C-reactive protein and erythrocyte sedimentation rate. Serum MIP-1alpha levels of the patients with FMF in acute attack period were significantly higher than the patients in remission and healthy controls (p=0.02 and p=0.038, respectively). MIP-1alpha levels were weakly correlated with CRP (r=0.32, p=0.032) levels. MIP-1alpha may have a role in the pathogenesis of FMF attacks. MIP-1alpha and other chemokines may constitute a link between the innate immune system and FMF.     

Association of severe noncerebral Plasmodium falciparum malaria in Brazil with expressed PfEMP1 DBL1 alpha sequences lacking cysteine residues

BACKGROUND: Cytoadherence and rosetting contribute to the development of severe Plasmodium falciparum malaria. In Brazil,severe falciparum malaria is mostly associated with renal or pulmonary complications and very rarely with cerebral malaria. The most N-terminal DBL1 alpha domain of PfEMP1, a protein encoded by the var multigene family mediates rosetting. We analyzed parasites of Brazilian patients with severe malaria to determine whether there were particular DBL1 alpha var sequences predominantly expressed in such patients. MATERIALS AND METHODS: DBL1 alpha var sequences were obtained from parasites of Brazilian patients with severe and mild malaria and were analyzed by standard bioinformatic programs. Three hundred twenty var DBL1 alpha sequences obtained from 80 Brazilian patients with mild malaria were spotted in high-density filters and hybridized to probes representing predominantly expressed sequences in parasites from patients with severe malaria. A DBL1 alpha domain was expressed in bacteria and used to demonstrate its binding capacity to erythrocytes by immunofluorescence. RESULTS: Forty-three different and unreported DBL1 alpha amino acid sequences were obtained. Sequences predominantly expressed in patients with severe malaria could be subgrouped due to deletions of 1-2-cysteine residues. These sequences were commonly found in the var gene repertoire of parasites from patients with mild malaria, yet they were rarely expressed in these patients. A recombinant protein representing the most abundantly expressed sequence detected in one patient with severe malaria bound directly to uninfected erythrocytes. CONCLUSIONS: This is the first report showing an association of severe noncerebral malaria from Brazil with particular DBL1 alpha sequences.     

Deoxyribonucleic acid (DNA) polymorphism of the alpha 1-antitrypsin gene in chronic lung disease.

Specific gene probes were used to study restriction fragment length polymorphisms of the human alpha 1-antitrypsin gene. A polymorphism due to loss of a recognition site for the restriction enzyme Taq I was identified in eight of 42 patients with bronchiectasis and nine of 49 patients with pulmonary emphysema, none of whom had alpha 1-antitrypsin deficiency. Among a control group without lung disease the polymorphism was significantly less frequent, being found in only five of 101 apparently healthy blood donors. The deoxyribonucleic acid (DNA) polymorphism was also present in two of 14 unrelated patients with alpha 1-antitrypsin deficiency, indicating a lack of association with any specific alpha 1-antitrypsin protein phenotype. The polymorphism identified in this study may be a new marker for genetic predisposition to chronic lung disease.     

Hydrochlorothiazide, allopurinol and magnesium oxide in the treatment of recurrent calcium urolithiasis]

The study aimed at presenting own experience in prevention of new urinary calculi in 18 patients with metabolically active calcium urolithiasis treated with hydrochlorothiazide in daily doses of 100 mg (group I) for 2 years, and in 6 patients with the same disease treated with magnesium oxide in daily doses 300 mg twice a day (group II) for average period of 10 months. In 9 patients a new calculus was formed during the treatment with hydrochlorothiazide, in 7 patients no recurrence was noted, and in 2 remaining patients the results were controversial (coral calculus). Therefore, patients were subdivided into group Ia (failure of hydrochlorothiazide therapy), and group Ib (no recurrence noted). Hydrochlorothiazide did not lead to the stable decrease in the saturation of the urine with calcium oxalate in group Ia whereas in group Ib (without recurrence of urolithiasis) the content of calcium oxalate in the urine was significantly lower than that in group Ia after a 2-year treatment with hydrochlorothiazide (p < 0.01) Recurrence of the disease was seen only in one patient of group II, i.e. treated with magnesium oxide. The treatment of the recurrent calcium urolithiasis is justified and efficient in those patients in whom therapy decreases the content of calcium oxalate in the urine.     

Diversity in Protein Profiles of Individual Calcium Oxalate Kidney Stones

Calcium oxalate kidney stones contain low amounts of proteins, some of which have been implicated in progression or prevention of kidney stone formation. To gain insights into the pathophysiology of urolithiasis, we have characterized protein components of calcium oxalate kidney stones by proteomic approaches. Proteins extracted from kidney stones showed highly heterogeneous migration patterns in gel electrophoresis as reported. This was likely to be mainly due to proteolytic degradation and protein-protein crosslinking of Tamm-Horsfall protein and prothrombin. Protein profiles of calcium oxalate kidney stones were obtained by in-solution protease digestion followed by nanoLC-MALDI-tandem mass spectrometry, which resulted in identification of a total of 92 proteins in stones from 9 urolithiasis patients. Further analysis showed that protein species and their relative amounts were highly variable among individual stones. Although proteins such as prothrombin, osteopontin, calgranulin A and calgranulin B were found in most stones tested, some samples had high contents of prothrombin and osteopontin, while others had high contents of calgranulins. In addition, calgranulin-rich stones had various neutrophil-enriched proteins such as myeloperoxidase and lactotransferrin. These proteomic profiles of individual kidney stones suggest that multiple systems composed of different groups of proteins including leucocyte-derived ones are differently involved in pathogenesis of individual kidney stones depending on situations.     

The acute-phase response in endometriosis of women

Peritoneal fluid volume was determined and concentrations of C-reactive protein, alpha 1-antitrypsin, acid-alpha 1-glycoprotein, alpha 2-macroglobulin, haptoglobin, complement factors C3 and C4, IgG, IgA and IgM were measured in the supernatant of the peritoneal fluid and in serum by means of a radial-immunodiffusion technique in 25 patients with and in 45 patients without endometriosis. Peritoneal fluid volume was not different between the two groups. The peritoneal fluid:serum ratios for the proteins determined showed a significant inverse correlation with their molecular weight in both groups, indicating that their presence in peritoneal fluid is governed by exudation according to their molecular weight, rather than by active production in, or selective release into, the peritoneal cavity. In control patients only, the ratios of most of the individual proteins studied were significantly higher in the luteal than in the follicular phase. We suggest that the high values of peritoneal fluid:serum ratios in endometriotic tissue and peritoneal macrophages. In the luteal phase, the cycle-dependent increase of protein exudation obscures this additional contribution. We conclude that endometriosis does not cause marked intra-abdominal inflammatory changes. If the presence of endometriosis lowers fecundity, the mechanism probably does not involve acute-phase protein synthesis.     

Sequence and acute phase regulation of rat alpha 1-inhibitor III messenger RNA

cDNA clones coding for the plasma proteinase inhibitor alpha 1-inhibitor III were isolated from an acute phase rat liver library. The isolates could be divided into four groups with characteristic BamHI restriction fragment patterns. The identity of the prototype clone pRLA1I3/2J was established by comparison with the published amino acid sequence of the purified protein. It codes for a 1477-amino acid precursor polypeptide with a 24-residue signal peptide. The mature protein shares 58% overall sequence identity with rat alpha 2-macroglobulin and contains a typical internal thiolester sequence. Twenty-two of its twenty-three cysteinyl residues are conserved with alpha 2-macroglobulin implying similar tertiary structure. However, the prototype alpha 1-inhibitor III sequence differed significantly from the rat and human alpha 2-macroglobulin sequences in its bait region suggesting alpha 1-inhibitor III possesses proteinase inhibitory specificities different from those of alpha 2-macroglobulin. The variant alpha 1-inhibitor III clone pRLA1I3/2J from a second cDNA group also differed from the prototype in the bait region coding sequence, although both specify similar signal peptides and NH2 termini. The observation of variant cDNA classes suggests that acute phase rat livers produce a heterogeneous mixture of alpha 1-inhibitor III mRNA molecules. Evidence was obtained for the presence of at least four different alpha 1-inhibitor III-related genes in the rat genome. During the first 24 h of an acute phase response the abundance of hepatic alpha 1-inhibitor III mRNA was decreased 3-4-fold. This decrease was of the same order of magnitude as the reported reduction of the corresponding plasma protein concentration, suggesting that in the early phase of the acute inflammatory response the plasma concentration of this protein is mainly controlled through the abundance of its hepatic mRNA.     

The hinge region of the scaffolding protein of cell contacts, zonula occludens protein 1, regulates interacting with various signaling proteins

Zonula occludens protein 1 (ZO-1) is a ubiquitous scaffolding protein, but it is unknown why it functions in very different cellular contacts. We hypothesized that a specific segment, the unique hinge region, can be bound by very different regulatory proteins. Using surface plasmon resonance spectroscopy and binding assays to peptide libraries, we show, for the first time, that the hinge region directly interacts with disparate signal elements such as G-proteins alpha 12 and alpha i2, the regulator of G-protein signaling 5, multifunctional signaling protein ahnak1, and L-type Ca2+-channel beta-2-subunit. The novel binding proteins specifically bound to a coiled coil-helix predicted in the hinge region of ZO-. The interactions were modulated by phosphorylation in the hinge helix. Activation of the G-proteins influenced their association to ZO-1. In colon cells, G alpha i2 and ZO-1 were associated, as shown by coimmunoprecipitation. After cotransfection in kidney cells, G alpha i2 barely colocalized with ZO-1; the colocalization coefficient was significantly increased when epinephrine activated G-protein signaling. In conclusion, proteins with different regulatory potential adhere to and influence cellular functions of ZO-proteins, and the interactions can be modulated via its hinge region and/or the binding proteins.     

Demonstration of serum alpha1-macroglobulin on leucocyte surfaces in non-pregnant and pregnant rats

alpha1-macroglobulin (alpha1-M) was demonstrated by indirect immunofluorescence staining on the surface membrane of leucocytes obtained from rat blood and lymphoid tissue. The incidence of alpha1-M positive cells correlated with serum levels of the protein. Compared with blood, a significantly higher percentage of staining cells was demonstrated in spleen and lumbar lymph nodes. During pregnancy, when serum levels of alpha1-M were significantly elevated, the proportion of alpha1-M staining leucocytes increased in blood, Thymus and lymph nodes but the incidence of alpha1-M positive spleen cells was unaffected. The comparatively low incidence of alpha1-M staining thymocytes, together with the presence of alpha1-M on glass-adherent cells and on leucocytes from congenitally athymic rats, suggest that this protein is associated predominantly with B cells and monocytes.     

The in vitro interactions of rat pancreatic elastase and normal and inflammatory ray serum.

The partition of labelled rat pancreatic elastase (EC 3.4.21.11) between the different protease inhibitors of rat plasma was studied at different levels of saturation of the inhibitors of rat plasma was studied at different levels of saturation of the inhibitor capacity of plasma with the enzyme. The reaction mixtures were analysed by immunoelectrophoretic methods utilizing specific antisera against the different inhibitors and by gel filtration on Sephadex G-200. Rat serum was shown to contain four elastase binding proteins. alpha 1-antitrypsin, alpha 1-macroglobulin and alpha 2-acute phase protein and alpha 1-inhibitor 3 which exhibits immunologic cross-reaction with human inter-alpha-trypsin inhibitor and is of similar molecular weight. With minute amounts of labelled elastase the partition among the binding protein was alpha 1-macroglobulin 60%, alpha 1-antitrypsin 24% and alpha 1-I3 16%. The 60% value of alpha 1-M bound radioactivity in normal serum corresponds to the sum of alpha 1-M and alpha 2-AP labelling in inflammatory serum.