Extensive characterizations of bacteria isolated from catheterized urine and stone matrices in patients with nephrolithiasis.

BACKGROUND: Urinary tract infections are generally known to be associated with nephrolithiasis, particularly struvite stone, in which the most common microbe found is urea-splitting bacterium, i.e. Proteus mirabilis. However, our observation indicated that it might not be the case of stone formers in Thailand. We therefore extensively characterized microorganisms associated with all types of kidney stones. METHODS: A total of 100 kidney stone formers (59 males and 41 females) admitted for elective percutaneous nephrolithotomy were recruited and microorganisms isolated from catheterized urine and cortex and nidus of their stones were analyzed. RESULTS: From 100 stone formers recruited, 36 cases had a total of 45 bacterial isolates cultivated from their catheterized urine and/or stone matrices. Among these 36 cases, chemical analysis by Fourier-transformed infrared spectroscopy revealed that 8 had the previously classified 'infection-induced stones', whereas the other 28 cases had the previously classified 'metabolic stones'. Calcium oxalate (in either pure or mixed form) was the most common and found in 64 and 75% of the stone formers with and without bacterial isolates, respectively. Escherichia coli was the most common bacterium (approximately one-third of all bacterial isolates) found in urine and stone matrices (both nidus and periphery). Linear regression analysis showed significant correlation (r = 0.860, P < 0.001) between bacterial types in urine and stone matrices. Multidrug resistance was frequently found in these isolated bacteria. Moreover, urea test revealed that only 31% were urea-splitting bacteria, whereas the majority (69%) had negative urea test. CONCLUSIONS: Our data indicate that microorganisms are associated with almost all chemical types of kidney stones and urea-splitting bacteria are not the major causative microorganisms found in urine and stone matrices of the stone formers in Thailand. These data may lead to rethinking and a new roadmap for future research regarding the role of microorganisms in kidney stone formation.

Effects of Orthosiphon grandiflorus, Hibiscus sabdariffa and Phyllanthus amarus extracts on risk factors for urinary calcium oxalate stones in rats.

PURPOSE: We evaluated the antilithic effect of Orthosiphon grandiflorus, Hibiscus sabdariffa and Phyllanthus amarus extracts on known risk factors for calcium oxalate stones in rats. MATERIALS AND METHODS: We divided 30 male Wistar rats into 5 equal groups. Controls were fed a standard diet and the remaining groups received a 3% glycolate diet for 4 weeks to induce hyperoxaluria. One glycolate fed group served as the untreated group and the others were given oral extracts of Orthosiphon grandiflorus, Hibiscus sabdariffa or Phyllanthus amarus at a dose of 3.5 mg daily. We collected 24-hour urine and blood samples. Kidneys were harvested for histological examination. We measured the renal tissue content of calcium and oxalate. RESULTS: The Hibiscus sabdariffa group showed significantly decreased serum oxalate and glycolate, and higher oxalate urinary excretion. The Phyllanthus amarus group showed significantly increased urinary citrate vs the untreated group. Histological examination revealed less CaOx crystal deposition in the kidneys of Hibiscus sabdariffa and Phyllanthus amarus treated rats than in untreated rats. Those rats also had significantly lower renal tissue calcium content than untreated rats. All parameters in the Orthosiphon grandiflorus treated group were comparable to those in the untreated group. CONCLUSIONS: Hibiscus sabdariffa and Phyllanthus amarus decreased calcium crystal deposition in the kidneys. The antilithic effect of Hibiscus sabdariffa may be related to decreased oxalate retention in the kidney and more excretion into urine while that of Phyllanthus amarus may depend on increased urinary citrate. In contrast, administering Orthosiphon grandiflorus had no antilithic effect.

Orthosiphon versus placebo in nephrolithiasis with multiple chronic complaints: a randomized control trial.

Nephrolithiasis in the communities of Northeast Thailand frequently presents with multiple chronic health complaints, i.e. myofascial pain, back pain, dyspepsia, arthralgia, headache, fatigue, frank paresthesia, dysuria and any of these aggravated by purine-rich food (PRF). We assessed the efficacy of Orthosiphon in treating subjects with at least two active symptoms and negative for urine white blood cells. Subjects were randomly allocated to two groups. Crude extract of Orthosiphon given in a capsule (equivalent to 1.6-1.8 g of dried leaves of Orthosiphon) two times a day to Group 1 (n = 36) and a placebo to Group 2 (n = 40) for 14 days. The medication for each subject was packed and its code kept secret until the data analysis. Both groups were asked not to consume any of 25 purine-rich foods (PRFs) during treatment. The primary measure was the reduced sum of active severity symptoms as recorded using the visual analog scale before and after therapy (i.e. on day 7 and 14). The data on 76 subjects were processed. The mean of the total scores (95% CI) of the symptoms in each group were decreased significantly (P < 0.001); 185.6 (153.3, 218.0) to 94.7 (58.2, 131.2) in the Orthosiphon group and 196.1 (164.4, 227.8) to 89.6 (62.8, 116.5) in the placebo group. When comparing between groups, no statistically significant difference was found. The mean consumption in PRFs was significantly decreased (P < 0.001) in both groups; however, Orthosiphon did not have additional benefit over placebo at 7 and 14 days of treatment during which they reduced these foods.

Uricosuric effect of Roselle (Hibiscus sabdariffa) in normal and renal-stone former subjects.

AIM OF THE STUDY: The Roselle (Hibiscus sabdariffa) was investigated for its uricosuric effect. MATERIALS AND METHODS: A human model with nine subjects with no history of renal stones (non-renal stone, NS) and nine with a history of renal stones (RS) was used in this study. A cup of tea made from 1.5 g of dry Roselle calyces was provided to subjects twice daily (morning and evening) for 15 days. A clotted blood and two consecutive 24-h urine samples were collected from each subject three times: (1) at baseline (control); (2) on days 14 and 15 during the tea drinking period; and (3) 15 days after the tea drinking was stopped (washout). Serum and 24-h urinary samples were analyzed for uric acid and other chemical compositions related to urinary stone risk factors. RESULTS: All analyzed serum parameters were within normal ranges and similar; between the two groups of subjects and among the three periods. Vis-à-vis the urinary parameters, most of the baseline values for both groups were similar. After taking the tea, the trend was an increase in oxalate and citrate in both groups and uric acid excretion and clearance in the NS group. In the RS group, both uric acid excretion and clearance were significantly increased (p<0.01). When the fractional excretion of uric acid (FEUa) was calculated, the values were clearly increased in both the NS and SF groups after the intake of tea and returned to baseline values in the washout period. These changes were more clearly observed when the data for each subject was presented individually. CONCLUSIONS: Our data demonstrate a uricosuric effect of Roselle calyces. Since the various chemical constituents in Roselle calyces have been identified, the one(s) exerting this uricosuric effect need to be identified.

Chemical components of urinary stones according to age and sex of adult patients.

OBJECTIVE: Report the relationship of composition with age and sex of the patients. MATERIAL AND METHOD: A series of 426 urinary stones, 33 from the lower (LUT) and 393 from the upper urinary tract (UUT) of adults, were analyzed for their chemical composition using infrared spectroscopy. The majority of LUT stones were from males (n = 26) and in the age group beyond 60 years (n = 20). RESULTS: Calcium oxalate (CaOx) and uric acid and urate (UA-UR) were the main constituents in LUT stones of males and UA-UR and magnesium ammonium phosphate (MAP) of females. While UA-UR was distributed in all age group of males, it was only detected in elderly females. In cases of UUT stones, the peak finding for both sexes was for the 50-59-year-olds (age class). The MAP component was found more commonly in UUT stones of females, particularly in the younger age groups. CaOx and calcium phosphate (CaP) were the main components of UUT stones in both sexes (CaP was slightly more common in females) with the highest proportion in the 30-49-year-olds (age class), thereafter they declined and were replaced with UA-UR. CONCLUSION: Although the proportion of LUT stones in the present study was small, the present findings agree with previous studies on the role of both age and sex in the etiopathogeny of urinary stones.

Urinary citrate excretion in patients with renal stone: roles of leucocyte ATP citrate lyase activity and potassium salts therapy.

BACKGROUND: Hypocitraturia is a major metabolic abnormality in rural Northeast Thais with renal stones. These people also have low serum and urinary potassium and consume a high carbohydrate and low fat diet, which together might influence the intracellular metabolism and urinary excretion of citrate. METHODS: In Study A, we measured plasma and urinary chemistries and assayed leucocyte ATP citrate lyase (ACL) activity in 30 normal urban control subjects (Group A1) and 30 rural renal stone patients (Group A2) in Northeast Thailand. Some of the subjects from both groups were also used to evaluate the intake of carbohydrate, protein and fat. In Study B, we examined the effects of potassium salts therapy with another group of 30 rural renal stone patients: Group B1 (n = 15) treated with potassium chloride and Group B2 (n = 15) with potassium-sodium citrate (with an aim to achieve 42 mEq potassium, 21 mEq sodium and 62 mEq citrate per day for 1 month). RESULTS: In Study A, the leucocyte ACL activity of Group A1 was much lower than that of Group A2 (3.2 +/- 0.7 vs. 9.3 +/- 3.8 micromol acetylhydroxamate/mg protein/30 min, p < 0.0001). The plasma potassium, urinary excretions of potassium and citrate in Group A1 were higher than in Group A2. When data of the two groups were combined, urinary citrate excretion was inversely correlated with leucocyte ACL activity (r = 0.6783, p < 0.001). While the dietary protein intake did not differ between Groups A1 and A2, the carbohydrate intake by Group A1 was significantly lower (65.2 +/- 7.9% vs. 83.1 +/- 2.9%, p < 0.01) and fat higher (21.0 +/- 6.4% vs. 6.2 +/- 4.1%, p < 0.002) than Group A2. After treatment with potassium chloride (Group B1), only the potassium was increased (p < 0.001), while those treated with potassium-sodium citrate (Group B2) experienced a significant increase in urinary pH (p < 0.002), potassium (p < 0.001) and citrate (p < 0.001), and a decrease in leucocyte ACL activity (p < 0.001). CONCLUSIONS: Compared to normal subjects, renal stone patients have low urinary citrate excretion with high leucocyte ACL activity. In Northeast Thailand, low potassium status and a high carbohydrate and low fat diet may cause the increased ACL activity. However, hypokaliuria, hypocitraturia and high leucocyte ACL activity can be corrected by potassium-sodium citrate salt therapy.

In vivo 31P-MRS assessment of muscle-pH, cytolsolic-[Mg2+] and phosphorylation potential after supplementing hypokaliuric renal stone patients with potassium and magnesium salts.

Renal stone patients in rural northeast Thailand have a low potassium and magnesium status and low urinary excretion of citrate. We measured the changes of urinary citrate excretion and assessed in vivo skeletal muscle metabolism for intracellular-pH, cytosolic-[Mg(2+)] and phosphorylation potential (using the phosphorus magnetic resonance spectroscopy (31)P-MRS) after oral supplementation to hypokaliuric renal stone patients with oral potassium and magnesium salts. The patients comprised four groups: Group 1 (n = 10) control, Group 2 (n = 3), Group 3 (n = 5) and Group 4 (n = 6) supplemented for a month with potassium citrate, potassium citrate plus amino acid chelated magnesium, and potassium-magnesium citrate, respectively. Though urinary citrate excretion was increased in all three supplemented groups, the increases in intracellular-pH, cytosolic-[Mg(2+)] and phosphocreatine (PCr)/beta-ATP were prominent only in Group 3. The increase in PCr/beta-ATP was also observed in Group 4.

Proton magnetic resonance spectroscopy of the kidney in renal stone disease.

Previous studies of renal stone disease (RSD) in Thailand indicated abnormal urinary aggregator and inhibitor composition among farmers with excessive sweat loss. Our aim was to compare the proton MR spectra obtained from the kidneys of 32 proven cases of RSD (aged 38 to 65 yrs) with nine age-matched normal control subjects. We used the STEAM sequence with TE = 15 ms and TR = 2,000 ms. The spectra at 3.25, 3.6 and 3.9 ppm were analyzed. The results showed a correlation between the three peaks (p < 0.001), however, there was no significant difference between the RSD group and the normal control subjects. We therefore concluded that there was no overloading of these osmolytes among the renal stone patients.

Magnesium and zinc status in survivors of sudden unexplained death syndrome in northeast Thailand.

Sudden Unexplained Death Syndrome (SUDS) is a major health problem in rural residents of Northeast Thailand. The cause of death in SUDS is suspected to be cardiovascular abnormalities. As magnesium (Mg) and zinc (Zn) deficiency contribute significantly to several cardiovascular diseases, we investigated the Mg- and Zn-status of patients with sudden respiratory distress and cardiac arrest who had survived resuscitation attempts or a near-SUDS episode (N-SUDS). The following subjects were enrolled: 12 N-SUDS inhabitants of rural Northeast Thailand (rural group 1, R1), 13 rural villagers with no past history of N-SUDS (rural group 2, R2), 15 urban Northeasterners (urban group 1, U1); 13 Bangkokians (urban group 2, U2). All subjects were free of structural heart disease. Magnesium and zinc were assessed by atomic absorption spectrophotometry of samples of plasma, red blood cells (RBC), white blood cells (WBC), and 24-hour urine. The mean levels of magnesium in the RBC, WBC, and 24-hour urine of N-SUDS patients (R1) were significantly lower than those of the urban groups (U1 and U2), while the plasma levels did not show any differences. When comparing the Zn-status of R1 with that of the urban groups (U1 and U2), the plasma, RBC, and WBC levels were found to be significantly lower in R1 (except for the RBC-Zn of the U1 group), while the 24-hour urine levels was higher. Although the magnesium and zinc parameters were not significantly different between the rural groups R1 and R2, the prevalence of hypomagnesuria (<2.2 mmol/day), hypozincemia (<9.7 micromol/l), and hyperzincuria (>10.7 micromol/day) was higher in the R1 group. These findings suggest that the homeostasis of both magnesium and zinc is altered in N-SUDS patients. Similar alterations, to a lesser degree, were observed in those people living in the same rural environment (R2).

Changes in erythrocyte contents of potassium, sodium and magnesium and Na, K-pump activity after the administration of potassium and magnesium salts.

Low potassium and magnesium status and decreased Na, K-pump activity is an endemic condition among rural Northeast Thais. The authors examined the effect of supplementing potassium and magnesium on erythrocyte potassium, sodium and magnesium content and on Na, K-pump activity. Rural Northeast Thai renal stone patients (62) were recruited, divided into four groups and supplemented for one month with potassium chloride (Group1, n = 16), potassium-sodium citrate (Group2, n = 15), chelated magnesium (Group 3, n =16) and potassium-magnesium citrate (Group 4, n =15) in order to achieve 40 mmol potassium, 10 mmol magnesium and 60 mmol citrate daily. After supplementation with potassium (Groups 1, 2 and 4), plasma potassium and Na, K-pump activity rose significantly in Groups 1, 2 and 4, but erythrocyte potassium rose only in Groups 2 and 4. When supplementing elemental magnesium (Groups 3 and 4), the chelated magnesium caused a significant increase in plasma potassium, erythrocyte potassium, sodium and magnesium without a significant increase in Na, K-pump activity. By contrast, potassium-magnesium citrate caused a significant increase in erythrocyte potassium and magnesium and Na, K-pump activity, but depressed erythrocyte sodium. These results suggest the forms of potassium and /or magnesium salts being supplemented should be considered because they affect erythrocyte potassium, sodium and magnesium content and Na, K-pump activity differently.

The effects of potassium and magnesium supplementations on urinary risk factors of renal stone patients.

The effects of potassium and magnesium supplementation on urinary risk factors for renal stone disease were studied in 61 renal stone patients. The subjects were divided into four groups and supplemented for a period of one month with potassium chloride (KCl, Group 1), potassium sodium citrate (K Na citrate, Group 2), magnesium glycine (Mg glycine, Group 3) and potassium magnesium citrate (K Mg citrate, Group 4) with a daily dose of 42 mEq potassium, 21 mEq magnesium or sodium and 63 mEq citrate, accordingly. The results showed that serum potassium and magnesium of all four groups normalized after the supplementation. Though urinary potassium significantly increased in all three groups supplemented with elemental potassium containing solutions [i.e. KCl (p < 0.001), K Na citrate (p < 0.001) and K Mg citrate (p < 0.001)] only K Na citrate and K Mg citrate, caused a significant increase in urinary pH and citrate but decrease in calcium. Supplementation with Mg glycine in Group 3 although caused a significant increase in urinary magnesium, its effects on urinary pH, citrate and calcium, however, were similar to KCl, in that they caused a significant decrease in urinary pH without any change in urinary citrate or calcium. Supplementation with K Mg citrate in Group 4 seems to have given the best results, as far as lowering stone risk factors in that it caused an increase in urinary pH, potassium and citrate and decreased calcium excretions similar to K Na citrate in Group 2. In addition, K Mg citrate also caused the enrichment of urine with magnesium, another inhibitor of calcium-containing stones. Although the four supplements had no effect on urinary saturation of calcium oxalate salt, their effects on the saturations of brushite (CaHPO4 x 2H2O), octacalcium phosphate (Ca8H2 (PO4)6 x 5H2O) and uric acid were clearly associated with changes in urinary pH. Therefore, in Group 1 and 3, subjects having a decrease in urinary pH, also experienced a significant increase in uric acid saturation. Though the saturation of brushite and octacalcium phosphate in Group 2 and 4 and the sodium acid urate in Group 2 were significantly increased, these urinary risk factors could be overcome, however, by the concomitant increase in urinary citrate. The present results demonstrate that for those stone vulnerable subjects having a high risk of potassium and magnesium depletion, to obtain the best therapeutic results, they should be provided supplementations of both potassium and magnesium together and also in the forms that would result in the delivery of an alkali loading effect.

Metabolic enzymes, antioxidants, and cytoskeletal proteins are significantly altered in vastus lateralis muscle of K-depleted cadaveric subjects.

Molecular mechanisms underlying myopathy caused by prolonged potassium (K) depletion remain poorly understood. In the present study, we examined proteome profile of vastus lateralis muscle obtained from cadaveric subjects who had K depletion (KD) (muscle K<80 micromol/g wet weight) compared to those who had no KD (NKD) (muscle K>or=80 micromol/g wet weight) (n=6 per group). Muscle proteins were extracted, resolved by 2-DE, and visualized with CBB-R250 stain. Spot matching and intensity analysis revealed significant changes in levels of 11 (6 increased and 5 decreased) protein spots in the KD group. Q-TOF MS and MS/MS analyses identified these altered proteins as metabolic enzymes (aldehyde dehydrogenase 1A1, uridine diphosphoglucose pyrophosphorylase, enolase 1, cytosolic malate dehydrogenase, and carbonic anhydrase III), antioxidants (peroxiredoxin-3 isoform b), cytoskeletal proteins (slow-twitch skeletal troponin I and myosin light chain 2), and others. These altered proteins are involved in many cellular functions, including bioenergetics, acid-base regulation, oxidative stress response, and muscle contractility. Validation was done by Western blot analysis, which confirmed the increased level of peroxiredoxin-3 and decreased level of troponin-I in the KD muscle. Linear regression analysis also revealed a significant negative correlation between peroxiredoxin-3 level and muscle K content (r=-0.887; p<0.001), as well as a significant positive correlation between troponin-I level and muscle K content (r=0.618; p<0.05). Our results implicate the important roles these altered proteins play in the development of KD-associated myopathy.

Serial analyses of postmortem changes in human skeletal muscle: A case study of alterations in proteome profile, histology, electrolyte contents, water composition, and enzyme activity.

Postmortem tissues are frequently used in forensic investigation, clinical studies, and biomedical research. It is well known that the shorter period from death to analyses provides the more accurate results. However, the longest postmortem interval that still provides the reliable data remains unclear. We performed serial analyses of postmortem changes in proteome profile, histology, electrolyte contents, water composition, and enzyme activity in human vastus lateralis muscle from a male cadaver (died from a motorcycle accident). This uninjured muscle was sectioned into several 1-cm(3 ) cubes and stored in individual closed tubes at 4 or 25°C for 0, 2, 4, 6, 12, 24 or 48 h prior to proteomic, histological, chemical and biochemical analyses. At 4°C, the 2-DE proteome profile remained unchanged until 24 h, when some poorly focused protein spots and significant decrease in the total number of visualized spots were observed. These changes were detectable earlier (12 h) in the samples stored at 25°C. Profound vacuolization and autolysis started at 24 and 6 h for the samples stored at 4ºC and 25°C, respectively. K and Mg contents began to increase at 12 and 48 h, respectively, for both temperatures. However, the increase in Na and Ca contents began at 24 h in the samples stored at 4°C, but started earlier (12 h) in those stored at 25°C. Water content started to decline at 48 and 24 h in the samples stored at 4 and 25°C, respectively. Muscle lactate dehydrogenase activity began to be out of range at 12 h for both temperatures. These findings demonstrate that storing the samples at 4°C could delay some of the aforementioned changes, which occurred more rapidly at 25°C. Our results also suggest that muscle proteome profile, histology, electrolyte contents, water composition, and enzyme activity should be analyzed within the optimal postmortem intervals, which vary among individual analyses, to obtain the most reliable data.

The levels of lycopene, alpha-tocopherol and a marker of oxidative stress in healthy northeast Thai elderly.

An imbalance between oxidative stress and antioxidant capacity has been proposed to play an important role in the development and progression of chronic diseases in the elderly. The present study was carried out to investigate correlation between the serum antioxidants (lycopene and alpha-tocopherol) and malondialdehyde (MDA), a marker of oxidative stress in the healthy Thai elderly. The 207 healthy subjects aged 60-91 years old (72 males and 135 females) in Khon Kaen province, Thailand were enrolled in this study. They were interviewed by questionnaires about smoking habit. Serum lycopene and alpha-tocopherol levels were determined by high performance liquid chromatography (HPLC). MDA was measured by thiobarbituric assay. Serum lycopene and alpha-tocopherol levels in the elderly were 0.27 micromol/L (95% CI = 0.23-0.31) and 22.10 micromol/L (95% CI = 20.99-23.22), respectively. Males had significant lower serum lycopene and alpha-tocopherol levels than females (p<0.001). Of 72 males, 31.94% are current smokers whereas 1.4% of 135 females are current smokers. Current smokers had significantly lower serum lycopene (0.17 +/- 0.11 micromol/L) than current non-smokers (0.28 +/- 0.27 micromol/L) (p=0.0439) but level of alpha-tocopherol had non significance (p=0.210). Moreover, the current smokers had higher MDA malondialdehyde level (1.55 +/- 0.10 micromol/L) than the current non-smokers (1.35 +/- 0.04 micromol/L) (p=0.094). Thus, dietary antioxidant supplementation from local fruits and vegetables may have a beneficial role in the prevention of chronic diseases at high-risk oxidative stress such as smoking in these elderly.

K+, Na+, Mg2+, Ca2+, and water contents in human skeletal muscle: correlations among these monovalent and divalent cations and their alterations in K+ -depleted subjects.

None of previous studies had simultaneously analyzed the K(+), Na(+), Mg(2+), and Ca(2+) contents in human skeletal muscle. We examined extensively and simultaneously the levels of all these cations and examined water content in vastus lateralis and pectoralis major muscles in 30 northeastern Thai men who were apparently healthy but died from an accident. Specimen collection was performed within 6 h of death. We used atomic absorption or flame photometry to measure the level of muscle cation. Histopathology of muscle and kidney was also evaluated. K(+), Na(+), Mg(2+), and Ca(2+) contents in vastus lateralis were 84.74 +/- 1.50, 38.64 +/- 0.77, 7.58 +/- 0.17, and 0.94 +/- 0.06 micromol/g wet weight, respectively, whereas K(+), Na(+), and Mg(2+) contents in pectoralis major were 82.83 +/- 1.54, 37.57 +/- 0.72, and 7.30 +/- 0.17 micromol/g wet weight, respectively. The water component was comparable in vastus lateralis and pectoralis major (78.66 +/- 0.41 and 78.09 +/- 0.56 %, respectively). Based on muscle K(+) levels, we divided the subjects into 2 main groups: K(+)-depleted (KD) group (K(+) < 80 micromol/g wet weight; n = 7) and non-K(+)-depleted (NKD) group (K(+) > or = 80 micromol/g wet weight; n = 23). In the KD muscle, Na(+) and Ca(2+) levels were significantly higher, whereas the level of Mg(2+) was significantly lower. Linear regression analysis showed significant correlations of K(+) and Mg(2+) levels and between Na(+) and Ca(2+). However, K(+) and Mg(2+) had the negative correlation with Na(+) and Ca(2+). Histopathologic examination showed no change in the KD muscles, whereas 29% (2 of 7) of the KD kidneys had vacuolization in proximal renal tubular cells. Our study not only provided the descriptive data but also implied the balance or homeostasis of these monovalent and divalent cations in their muscle pools.

Effects of potassium-magnesium citrate supplementation on cytosolic ATP citrate lyase and mitochondrial aconitase activity in leukocytes: a window on renal citrate metabolism.

BACKGROUND: An increase in urinary citrate excretion is associated with a decrease in activity of renal cortical cytosolic ATP citrate lyase (ACL) and mitochondrial aconitase (m-aconitase). Because potassium-magnesium citrate causes an increase in urinary citrate excretion, we decided to assess its effects on ACL and m-aconitase in the leukocytes of renal stone patients. METHODS: Twenty male renal stone patients were supplemented with potassium-magnesium citrate twice daily (i.e. 42 mEq potassium, 21 mEq magnesium, and 63 mEq citrate per day) for a period of 1 month. Two 24-h urine and one 15-mL heparinized blood samples were collected from each patient before and after supplementation. Urine samples were analyzed for relevant biochemical compositions. Leukocytes were separated from blood samples by centrifugation and assayed for ACL and m-aconitase activity. RESULTS: Supplementation with potassium-magnesium citrate significantly increased urinary pH (P < 0.005) and excretions of potassium (P < 0.001), magnesium (P < 0.001) and citrate (P < 0.0001). The activity of both ACL and m-aconitase were significantly decreased (P < 0.004 and P < 0.02 respectively). The decrease in ACL activity was inversely correlated with an increase in urinary excretion of both potassium (r = -0.620, P < 0.0001) and citrate (r = -0.451, P < 0.004). A similar inverse correlation was observed between m-aconitase activity and urinary excretion of citrate (r = -0.322, P < 0.043). CONCLUSION: Changes in enzyme activity, related to citrate metabolism in leukocytes, might reflect the status of renal tubular cells.

Renal tubular cell damage and oxidative stress in renal stone patients and the effect of potassium citrate treatment.

Our objective was to evaluate the oxidative stress and renal tubular cell damage in patients who have renal stones compared to normal subjects. The patients were re-evaluated after 1-months supplementation with potassium citrate. We recruited 30 patients (11 males and 19 females) diagnosed with kidney stones and scheduled for surgical stone removal the following month, and 30 healthy non-stone formers (14 males and 16 females). Two 24-h urine samples and one heparinized blood sample were collected from each subject. Plasma was separated from the erythrocytes and assayed for creatinine, potassium, sodium, calcium, magnesium, phosphate, malondialdehyde (MDA, a lipid peroxidation product) (P-MDA), protein thiol as an indicator of protein oxidation, and vitamin E. Erythrocytes were analysed for MDA (E-MDA), reduced glutathione (GSH) and cellular glutathione peroxidase (cGPx) activity. The urine was analyzed for pH, creatinine, potassium, sodium, calcium, magnesium, phosphate, oxalate, citrate, MDA (U-MDA), total protein (U-protein) and N-acetyl-beta-glucosaminidase (NAG) activity. For the stone patients, urine and blood samples were re-evaluated after supplementation with potassium citrate (60 mEq/day) for 1 month. Renal stone patients had higher plasma creatinine and lower plasma potassium, urinary pH, potassium, magnesium, phosphate and citrate than the controls. The patients had higher P-MDA, E-MDA, U-MDA, U-protein and NAG activity, but lower GSH, cGPx activity, protein thiol and vitamin E, when compared with controls. After potassium citrate supplementation, P-MDA and E-MDA decreased while plasma vitamin E, urinary NAG activity and citrate increased. Renal stone disease is associated with high oxidative stress and damage to renal tubular cells. These abnormalities are coincident with an increase in blood lipid peroxidation products and a decrease in antioxidant status. Although supplementation with potassium citrate improved urinary citrate levels and oxidative stress, it neither reduced urinary lipid peroxidation products nor remedied the damage to renal tubular cells, probably due to the existence of kidney stones.