Pathophysiological analysis of uninephrectomized db/db mice as a model of severe diabetic kidney disease.

Diabetic nephropathy, included in diabetic kidney disease (DKD), is the primary disease leading to end-stage renal disease (ESRD) or dialysis treatment, accounting for more than 40% of all patients with ESRD or receiving dialysis. Developing new therapeutics to prevent the transition to ESRD or dialysis treatment requires an understanding of the pathophysiology of DKD and an appropriate animal model for drug efficacy studies. In this study, we investigated the pathophysiology of diabetic kidney disease with type 2 diabetes in uninephrectomized db/db mice. In addition, the nephrectomized db /db mice from 10 weeks to 42 weeks were used to assess the efficacy of long-term administration of the angiotensin-II-receptor antagonist losartan. The blood and urinary biochemical parameters, main pharmacological endpoint of the losartan therapy, were periodically measured. And at the end, histopathological analysis was performed. Uninephrectomized db/db mice clearly developed obesity and hyperglycemia from young age. Furthermore, they showed renal pathophysiological changes, such as increased urinary albumin-creatinine ratio (UACR) (the peak value 3104 ± 986 in 40-week-old mice), glomerular hypertrophy and increased fibrotic areas in the tubulointerstitial tubules. The blood pressure in the losartan group was significantly low compared to the normotensive Vehicle group. However, as expected, Losartan suppressed the increase in UACR (829±500) indicating the medication was sufficient, but the histopathological abnormalities including tubular interstitial fibrosis did not improve. These results suggest that the uninephrectomized db/db mice are useful as an animal model of the severe DKD indicated by the comparison of the efficacy of losartan in this model with the efficacy of losartan in clinical practice.

ALKBH3, a human AlkB homologue, contributes to cell survival in human non-small-cell lung cancer.

BACKGROUND: We have demonstrated for the first time that a novel human AlkB homologue, ALKBH3, contributes to prostate cancer development, but its clinical and biological roles in lung cancer remain unclear. METHODS: Expression of both mRNA and protein of PCA-1 was examined by RT-PCR and western blotting. We also assessed association with senescence and in vivo ALKBH3 treatment on orthotopic tumour cell inoculation, and analysed it clinicopathologically. RESULTS: We have since found novel biological roles for ALKBH3 in human lung cancers, particularly in adenocarcinoma. Our immunohistochemical analysis of human adenocarcinomas and squamous cell carcinomas of the lung not only showed overexpression of ALKBH3 in these tumours but the percentage of cells positive for ALKBH3 also correlated statistically to recurrence-free survival in adenocarcinoma. Knockdown of ALKBH3 by siRNA transfection induced expression of p21(WAF1/Cip1) and p27(Kip1) in the human lung adenocarcinoma cell line A549, resulting in cell cycle arrest, senescence and strong suppression of cell growth in vitro. In vivo, peritoneal tumour growth and dissemination was inhibited in nude mice, previously inoculated with the A549 cell line, by intraperitoneal injection of ALKBH3 siRNA + atelocollagen, as demonstrated by the reduction in both number and diameter of tumours developing in the peritoneum. CONCLUSION: We suggest that ALKBH3 contributes significantly to cancer cell survival and may be a therapeutic target for human adenocarcinoma of the lung.

Prognostic significance of CD45RO+ memory T cells in renal cell carcinoma.

BACKGROUND: Memory T cells are well known to have a critical role for host defense in humans. However, their role in actual human cancer remains largely unknown. In this study, we tried to reveal the clinical importance of tumour-infiltrating CD45RO+ memory T cells in renal cell carcinoma (RCC). METHODS: We analysed 105 patients with RCC, who received radical or partial nephrectomy. Those were 65 in TNM stage I, 7 in stage II, 15 in stage III, and 18 in stage IV, respectively. CD45RO expression was evaluated by immunohistochemistry. CD4 and CD8 expressions were also systematically assessed in the same manner. RESULTS: Patients with higher TNM stage or high nuclear grade were found to have higher densities of CD45RO. Furthermore, CD45RO status was positively correlated with preoperative C-reactive protein level. In prognostic analysis, CD45RO+lo patients had a significantly better prognosis than CD45RO+hi patients. There was also a significant difference between CD4+lo and CD4+hi groups, whereas no significant difference was observed in CD8 T-cell status. Finally, multivariate analysis revealed that CD45RO+ status was the independent prognostic factor for patient overall survival. CONCLUSION: CD45RO+ memory T-cell status has a significant independent prognostic value, indicating that the adaptive immune response is functionally critical in human RCC.

Clinical importance of B7-H3 expression in human pancreatic cancer.

BACKGROUND: B7-H3 is a new member of the B7 ligand family and regulates T-cell responses in various conditions. However, the role of B7-H3 in tumour immunity is largely unknown. The purpose of this study was to evaluate the clinical significance of B7-H3 expression in human pancreatic cancer and the therapeutic potential for cancer immunotherapy. METHODS: We investigated B7-H3 expression in 59 patients with pancreatic cancer by immunohistochemistry and real-time PCR. Furthermore, we examined the anti-tumour effect of B7-H3-blocking monoclonal antibody in vivo in a murine pancreatic cancer model. RESULTS: Tumour-related B7-H3 expression was abundant in most human pancreatic cancer tissues and was significantly higher compared with that in non-cancer tissue or normal pancreas. Moreover, its expression was significantly more intense in cases with lymph node metastasis and advanced pathological stage. B7-H3 blockade promoted CD8(+) T-cell infiltration into the tumour and induced a substantial anti-tumour effect on murine pancreatic cancer. In addition, the combination of gemcitabine with B7-H3 blockade showed a synergistic anti-tumour effect without overt toxicity. CONCLUSION: Our data show for the first time that B7-H3 may have a critical role in pancreatic cancer and provide the rationale for developing a novel cancer immunotherapy against this fatal disease.

Isoflurane exerts a short-term but not a long-term preconditioning effect in neonatal rats exposed to a hypoxic-ischaemic neuronal injury.

BACKGROUND: Isoflurane has been shown to induce tolerance against ischaemic injury in adult rodents. Although the delayed preconditioning effect of isoflurane has been demonstrated in neonatal rat pups, the acute preconditioning effects of isoflurane remained undetermined. The present study was therefore conducted to evaluate the acute preconditioning efficacy of isoflurane in neonatal rats subjected to a hypoxic-ischaemic (HI) injury. METHODS: Post-natal day 7 pups were exposed to 1 or 2% isoflurane in oxygen for either 30, 60 or 90 min. Fifteen minutes after isoflurane exposure, the pups were subjected to an HI injury induced by left common carotid artery ligation and exposure to 8% oxygen for 2 h. Pups not exposed to isoflurane or not subjected to HI served as controls. Histopathologic injury to the cortex and hippocampus was evaluated 7 and 49 days after HI. RESULTS: Isoflurane 2% exposure for 60 or 90 min before HI induced tolerance in the hippocampus and the number of normal neurons in the CA1 sector 7 days after HI was significantly greater than in non-preconditioned animals. This protective efficacy of isoflurane preconditioning was not observed 49 days after HI. CONCLUSIONS: Exposure of 2% isoflurane for at least 60 min is required to induce tolerance against HI injury in rat pups. However, this neuroprotective efficacy results in only transient neuroprotection.

Impact of a left-lobe graft without modulation of portal flow in adult-to-adult living donor liver transplantation.

In adult-to-adult living donor liver transplantation (LDLT), left-lobe grafts can sometimes be small-for-size. Although attempts have been made to prevent graft overperfusion through modulation of portal inflow, the optimal portal venous circulation for a liver graft is still unclear. Hepatic hemodynamics were analyzed with reference to graft function and outcome in 19 consecutive adult-to-adult LDLTs using left-lobe grafts without modulation of graft portal inflow. Overall mean graft volume (GV) was 398 g, which was equivalent to 37.8% of the recipient standard liver volume (SV). The GV/SV ratio was less than 40% in 13 of the 19 recipients. Overall mean recipient portal vein flow (PVF) was much higher than the left PVF in the donors. The mean portal contribution to the graft was markedly increased to 89%. Average daily volume of ascites revealed a significant correlation with portal vein pressure, and not with PVF. When PVP exceeds 25 mmHg after transplantation, modulation of portal inflow might be required in order to improve the early postoperative outcome. Although the study population was small and contained several patients suffering from tumors or metabolic disease, all 19 patients made good progress and the 1-year graft and patient survival rate were 100%. A GV/SV ratio of less than 40% or PVF of more than 260 mL/min/100 g graft weight does not contraindicate transplantation, nor is it necessarily associated with a poor outcome. Left-lobe graft LDLT is still an important treatment option for adult patients.

Effect of continuous morphine infusion on hypoxic-ischaemic brain damage of neonatal rats.

BACKGROUND: Opioids are commonly administered to critically ill neonates and infants for general anaesthesia and sedation. However, the clinical safety of these drugs, especially the effects on hypoxic-ischaemic damage of the developing brain, has not been well investigated. The present study was therefore conducted to investigate the effects of continuous morphine infusion on brain damage after hypoxic-ischaemic insults in neonatal rats. METHODS: Seven-day-old Sprague-Dawley rats were subjected to left common carotid artery ligation followed by a 90-min exposure of 8% oxygen. The rats were administered morphine (0.1, 0.3 or 1 mg/kg/h) or saline continuously for 72 h using osmotic minipumps. Seven days later, the rats were weighed and their brains were morphologically categorized into groups based on the following grades: 0=normal, 1=mild atrophy, 2=moderate atrophy, 3=atrophy with cystic cavitation <3 mm and 4=cystic cavitation >3 mm. For histological assessment, the ratio of the surviving neurons (ipsilateral/contralateral) was calculated in the cornu ammonis fields, CA1 and CA3, and the dentate gyrus (DG). RESULTS: One week after recovery (P14), the rats in the 1 mg/kg/h group showed significantly poorer weight gain compared with the other groups. However, the morphological score of the brains and the ratio of the surviving neurons in the CA1, CA3 and DG were similar among the groups. CONCLUSION: Our results indicate that continuous administration of morphine does not worsen brain damage 7 days after hypoxic-ischaemic insults in neonatal rats.

Prostatic volume and volume-adjusted prostate-specific antigen as predictive parameters for prostate cancer patients with intermediate PSA levels.

The object of the study was to examine the usefulness of volume-adjusted prostate-specific antigen (PSA) parameters for prediction of prostate cancer in the patients with intermediate PSA levels. The subjects were 235 patients with intermediate PSA levels (range: 4.1-10.0 ng/ml) whose prostate volume (PV) and prostate transition zone volume (TZV) were evaluated between August 1996 and April 2004. PSA, PV, TZV, PSA density (PSAD) (PSA/PV) and PSA transition zone density (PSATZD) (PSA/TZV) were assessed with the receiver operating characteristic (ROC) curve and the area under the curve (AUC). Simple and multivariate logistic regression analyses were used to analyze the odds ratios of age, PSA, PSAD, PSATZD, PV, TZV, digital rectal examination (DRE) and transrectal ultrasonography (TRUS) findings. Fifty-five patients (23.4%) of 235 patients had biopsy-proven prostate cancer. The univariate analysis revealed significant differences in the mean values of age, PSAD, PSATZD, PV, TZV and DRE between the patients with cancer and the non-cancer patients. The ROC curve analysis revealed that PV, TZV, PSAD and PSATZD had significant predictive values as compared with that of PSA. However, there was no difference in AUC between them. The stepwise logistic regression analysis showed that the age, PV, PSATZD and DRE had significant predictive values, and that PSATZD had the most predictive power. In conclusion, both PSAD and PSATZD had significant predictive values in discriminating prostate cancer. Furthermore, the stepwise logistic regression analysis showed that PSATZD had the strongest predictive value.

Frontal cerebral blood flow change associated with infant-directed speech.

OBJECTIVE: To examine the auditory perception of maternal utterances by neonates using near-infrared spectroscopy (NIRS). METHODS: Twenty full-term, healthy neonates were included in this study. The neonates were tested in their cribs while they slept in a silent room. First, two probe holders were placed on the left and right sides of the forehead over the eyebrows using double-sided adhesive tape. The neonates were then exposed to auditory stimuli in the form of infant-directed speech (IDS) or adult-directed speech (ADS), sampled from each of the mothers, through an external auditory speaker. RESULTS: A 2 (stimulus: IDS and ADS) x 2 (recording site: channel 1 (right side) and channel 2 (left side)) analysis of variance for these relative oxygenated haemoglobin values showed that IDS (Mean = 0.25) increased brain function significantly (F = 3.51) more than ADS (Mean = -0.26). CONCLUSIONS: IDS significantly increased brain function compared with ADS. These results suggest that the emotional tone of maternal utterances could have a role in activating the brains of neonates to attend to the utterances, even while sleeping.

Trisodium nitriloacetate monohydrate: promoting effect in urinary bladder carcinogenesis in rats treated with N-butyl-N-(4-hydroxybutyl)nitrosamine.

The effect of trisodium nitrilotriacetate monohydrate [(Na3NTA X H2O) CAS: 18662-53-8] on development of urinary bladder tumors in rats initiated with N-butyl-N-(4-hydroxybutyl)nitrosamine [(BBN) CAS: 3817-11-6] was studied. Twenty-one male inbred W rats 6 weeks of age were given drinking water containing 500 ppm of BBN for 4 weeks and then put on diet containing 10,000 ppm of Na3NTA X H2O for 28 weeks. Na3NTA X H2O promoted the development of urinary bladder tumors in rats treated with BBN. The incidences of papilloma and transitional cell carcinomas in the urinary bladder were 90% (18/20) and 25% (4/20), respectively, in rats treated with BBN and then Na3NTA X H2O and 0 in those treated with BBN or Na3NTA X H2O alone. The incidence of papillary or nodular hyperplasia in week 32 was 100% (20/20) in rats treated with BBN and then Na3NTA X H2O and 61% (13/21) in rats treated with BBN only.

beta-Cyclodextrin: promoting effect on the development of renal tubular cell tumors in rats treated with N-ethyl-N-hydroxyethylnitrosamine.

Injection (sc) of beta-cyclodextrin (beta-C) increased the number and size of renal tubular cell tumors in inbred Wistar (W) rats treated with 1,000 ppm of N-ethyl-N-hydroxyethylnitrosamine (EHEN). The incidence of renal tumors at the end of the 32-week experiment was 50% in rats treated with 1,000 ppm EHEN for 2 weeks and 100% in rats treated with 1,000 ppm EHEN for 2 weeks and then given daily sc injections of beta-C for 1 week. The incidence of renal tumors more than 3 mm in diameter was 70% in rats treated with 1,000 ppm EHEN before beta-C but 0% in rats treated with EHEN alone. In addition, beta-C promoted the development of renal tumors in rats treated with 500 ppm EHEN, which is a subthreshold dose for renal tubular cell tumorigenesis. These results show that beta-C promotes EHEN-induced renal tubular cell tumorigenesis.

4,4'-Diaminodiphenylmethane: promoting effect on the development of thyroid tumors in rats treated with N-bis(2-hydroxypropyl)nitrosamine.

4,4'-Diaminodiphenylmethane (4,4'-methylenedianiline) (DDPM) promoted the development of thyroid tumors in rats treated with a subeffective dose of N-bis(2-hydroxypropyl)nitrosamine (2,2'-dihydroxy-N-nitrosodipropylamine) (DHPN) for thyroid tumorigenesis. Male inbred W rats were given a single ip injection of 280 mg DHPN/100 g body weight and fed diets with or without 1,000 ppm DDPM. Thyroid tumor incidences at the end of week 20 of the experiment were 90% (19/21) in rats given DHPN and then DDPM and 28% (6/21) in rats given DHPN alone. The incidence of thyroid cancers was 9.5% (2/21) in rats first given DHPN and then DDPM. Untreated rats and rats given DDPM alone had no thyroid tumors after 20 weeks. Incidences of kidney tumors were 38% (8/21) in rats given DHPN and then DDPM and 28% (6/21) in rats given DHPN alone. No tumors were found in the kidneys and lungs of rats given DDPM alone and in those of control rats. Treatment with DDPM alone slightly but not significantly decreased the serum concentrations of thyroxine and triiodothyronine; treatment with DHPN plus DDPM had no such effect.

Trisodium nitrilotriacetate monohydrate: promoting effects on the development of renal tubular cell tumors in rats treated with N-ethyl-N-hydroxyethylnitrosamine.

The effect of trisodium nitrilotriacetate monohydrate [N,N-bis(carboxymethyl)glycine trisodium salt] (Na3NTA X H2O) on development of renal tubular cell tumors induced with N-ethyl-N-hydroxyethylnitrosamine [CAS:13147-25-6; 2-(ethylnitrosamino)-ethanol] (EHEN) was studied. Six-week-old male inbred W rats were given a diet containing 1,000 ppm of EHEN for 2 weeks and then a diet containing a high (10,000 ppm) or low (500 ppm) concentration of Na3NTA X H2O for 30 weeks. The rats were killed during week 32. The higher concentration of Na3NTA X H2O enhanced the development of renal tubular cell tumors and increased the number and size of tumors in rats treated with EHEN, but the lower concentration of Na3NTA X H2O did not. The incidence of renal tubular cell tumors in week 32 was 33% in rats treated with 1,000 ppm EHEN for 2 weeks, 100% in rats treated with 1,000 ppm EHEN for 2 weeks plus high Na3NTA X H2O diet for 30 weeks, and 39% in rats treated with 1,000 ppm EHEN for 2 weeks and then given low Na3NTA X H2O diet for 30 weeks. Numbers of atypical cell foci per kidney area (No./cm2) were 17.0 +/- 7.6 in rats treated with EHEN and high Na3NTA X H2O, 7.3 +/- 2.2 in rats treated with EHEN and low Na3NTA X H2O, 3.7 +/- 1.4 in rats treated with EHEN alone, and 1.0 +/- 2.4 in rats treated with high Na3NTA X H2O diet alone. Atypical cell foci retained a tubular pattern and consisted of basophilic cells with a large nucleus or clear cells with a small nucleus.

DL-Serine: promoting activity on renal tumorigenesis by N-ethyl-N-hydroxyethylnitrosamine in rats.

Subcutaneous injection of DL-serine increased the number and size of renal tubular cell tumors in male W rats treated with 500 or 1,000 ppm N-ethyl-N-hydroxyethylnitrosamine [(EHEN) CAS: 13147-25-6, 2-(ethylnitrosamino)ethanol]. At the end of the 32-week experiment, the incidences of renal tumors were 95% in rats treated with 1,000 ppm EHEN for 2 weeks and then given three sc injections of DL-serine every 2 weeks, 33% in rats treated with 1,000 ppm EHEN for 2 weeks, and 28% in rats treated with 500 ppm EHEN for 2 weeks and then given three sc injections of DL-serine every 2 weeks. No renal tumors were found in rats treated with 500 ppm EHEN alone or given three sc injections of DL-serine alone every 2 weeks.

Sex differential and dose dependence of phenobarbital-promoting activity in N-bis(2-hydroxypropyl)nitrosamine-initiated thyroid tumorigenesis in rats.

Studies were made on the dose and sex dependence of thyroid tumor development in rats pretreated with N-bis(2-hydroxypropyl)nitrosamine (DHPN) followed by exposure to various doses of phenobarbital (PB). A direct dose-response relationship in induction of thyroid tumors was found in both male and female rats. Upon feeding the DHPN-treated rats with basal diet containing 20, 100, 500, and 2500 ppm of PB, the incidences of follicular adenoma were, respectively, 8, 45, 70, and 66% in male rats and 12, 17, 50, and 58% in female rats. Development of papillary adenomas in male rats was observed only at the higher doses of PB, at incidences of 12 and 20% for doses of 500 and 2500 ppm. Follicular carcinoma was also seen at higher doses of PB, at 16 and 12%, respectively, for the 500- and 2500-ppm groups. Neither follicular nor papillary carcinomas were induced in female rats; only a low incidence of papillary adenoma (4%) was observed with a PB concentration as high as 2500 ppm. A single injection of DHPN resulted in production of approximately 1 tumor/female rat and 2.5 tumors/male rat. DHPN combined with posttreatment with PB at doses up to 500 ppm did not increase tumor yield in female rats, whereas a 3-fold increase was observed in male rats for the 500-ppm-treated groups. When PB was increased to 2500 ppm a marked increase (8-fold) in tumor yield in male rats was observed, in contrast to a less than 3-fold increase in similarly treated female rats.

Frequent alterations of the p14(ARF) and p16(INK4a) genes in primary central nervous system lymphomas.

To elucidate the role of p53/p16(INK4a)/RB1 pathways in the tumorigenesis of primary central nervous system lymphomas (PCNSLs), we have analyzed p14(ARF), p16(INK4a), RB1, p21(Waf1), and p27(Kip1) status in a series of their 18 sporadic cases of diffuse large B-cell lymphoma, using methylation-specific PCR, differential PCR, and immunohistochemistry. Homozygous deletion or methylation of p14(ARF) was detected in 10 (56%) PCNSLs, and they were almost entirely deletions (except 1 case). A total of 11 (61%) PCNSLs demonstrated homozygous deletion (6 cases) or methylation (5 cases) of p16(INK4a). Six tumors showed both p14(ARF) and p16(INK4a) homozygous deletions. Hypermethylation of the RB1 and the p27(Kip1) promoter region was detected in 2 (11%) cases, whereas p21(Waf1) methylation was not detected in any. Immunohistochemistry revealed loss of p14(ARF) and p16(INK4a) expression in 10 (56%) samples, correlating with the gene status. Four cases showed independent negative immunoreactivity for pRB and p27(Kip1), and nearly one-half of cases (8 of 18; 44%) were characterized by lack of p21(Waf1) expression. These results indicate that inactivation of p14(ARF) and p16(INK4a) by either homozygous deletion or promoter hypermethylation represents an important molecular pathogenesis in PCNSLs. Hypermethylation of RB1, p21(Waf1), and p27(Kip1) appears to be of minor significance, these genes being independently methylated in PCNSLs.

Histopathological and biochemical analyses of transplantable renal adenocarcinoma in rats induced by N-ethyl-N-hydroxyethylnitrosamine.

Transplantable renal adenocarcinoma can be readily induced in Wistar strain male rats by initiation with N-ethyl-N-hydroxy-ethylnitrosamine followed by promotion with beta-cyclodextrin. The transplantability rates of the tumors by s.c. inoculation in newborn rats were 33 and 50%, respectively, for tumors of the first and second passages, and 100% for both third and fourth passages. The transplantability rates were affected by route of inoculation; rates of 50 and 100% were observed for s.c. and i.p. inoculations, respectively. The growth rate of tumors induced by i.p. inoculation was 3-fold higher than that induced by s.c. injection. Macroscopically, most of the tumors grew in the s.c. tissue of inoculation sites. However, invasive growth of tumors in spleen, liver, stomach, peritoneum, and intestine were seen in 50% of the animals inoculated i.p.; metastatic cancers to lung were seen in 16%. Histologically, the tumors were well-differentiated adenocarcinomas composed of uniform cells resembling kidney tubular cells and appeared to be derived from normal kidney tissues. A 5-fold decrease in gamma-glutamyl transferase activity in tumor tissues was found as compared with that of nontumorous kidney tissues. Electrophoretic analysis of cellular proteins in polyacrylamide gels revealed that tumor tissues exhibited five new polypeptides with molecular weights of 81,000, 64,000, 59,000, 50,000, and 36,000 which were either lacking or undetectable in the nontumourous area and control kidney. In addition, protein banding patterns of transplantable renal tumor appeared to be more heterogeneous than those of primary kidney tumor.

Establishment of estrogen receptor-positive transplantable rat thyroid tumor cell lines in vivo.

We established 17 transplantable rat thyroid tumor cell lines from the primary thyroid tumor of rats induced by N-bis(2-hydroxypropyl)nitrosamine. Among the 17 tumor cell lines established, only two of them (D1 and G1) were estrogen receptor (ER) positive. These two cell lines were characterized with respect to transplantability, histological features, ER contents and cellular localization, and expression of ER message. The ER contents, determined by dextran-coated charcoal assay, were 13.3 and 20.7 fmol/mg protein for D1 and G1 cell lines, respectively. Scatchard plot analysis indicates that the dissociation constants (Kd) were 0.17 and 0.4 nM, respectively, for D1 and G1 cell lines. Sucrose density centrifugation analysis detected a hormone-receptor complex which sedimented at the 4S region, characteristic for ER. Immunohistological staining revealed that the ER was localized in the nuclei. The presence of ER in D1 and G1 cell lines was further confirmed by reverse transcriptase-polymerase chain reaction to detect the ER mRNA. These results demonstrated that ER is expressed in some thyroid tumors. The ER-positive transplantable tumor cell lines are useful for studying the direct effect of estrogen on thyroid tumors in vitro and in vivo.

Crystallization and preliminary diffraction studies of recombinant human granulocyte-stimulating factor (KW2228).

Human granulocyte colony-stimulating factor (hG-CSF) specifically stimulates proliferation of neutrophils. Two crystal forms of a mutant of hG-CSF expressed in Escherichia coli have been obtained using the hanging drop vapour diffusion method. One form is triclinic, space group P1, with cell dimensions a = 37.3 A, b = 46.4 A, c = 47.7 A, alpha = 105.5 degrees, beta = 98.0 degrees and gamma = 109.4 degrees. The other is monoclinic, space group C2, with cell dimensions a = 82.0 A, b = 49.2 A, c = 49.4 A and beta = 113.9 degrees. Both crystal forms diffract beyond 2.0 A and are suitable for X-ray analysis.

Pentosidine and its deposition in renal tissue in renal transplantation.

BACKGROUND: Advanced glycation end products (AGEs) accumulate in lesions of arteriosclerosis, Alzheimer's disease, rheumatoid arthritis, diabetic retinopathy, and diabetic nephropathy. Among AGEs, chemical quantification and immunohistologic methods for pentosidine have been established. Free pentosidine-eliminated by renal excretion- is mainly affected by renal function. In this study, we measured concentrations of plasma free and total pentosidine and immunohistologically investigated kidney graft biopsy specimens in patients after renal transplantation to investigate the renal function, plasma free and total pentosidine, and its relationship with deposition in the renal tissue. PATIENTS AND METHODS: In 28 patients who underwent renal transplantation from 1996 to 2003, we measured the time course of plasma concentrations of free pentosidine, total pentosidine, and serum creatinine starting right after renal transplantation. Thirty-four graft biopsy specimens were immunohistologically investigated using anti-pentosidine antibody. Plasma free and total pentosidine, and serum creatinine were measured at the same time. RESULTS: Plasma free and total pentosidine were positively correlated with serum creatinine. Plasma free pentosidine and serum creatinine reached nadir values on day 34.2 +/- 14.2, when the blood concentrations were 5.1 +/- 1.6 pmol/mL and 1.7 +/- 0.7 mg/dL, respectively. Plasma total pentosidine reached a nadir on day 116.5 +/- 39.7 when the plasma concentration was 4.0 +/- 1.5 pmol/mg. We correlated the time required to reach the nadir of plasma free and total pentosidine concentrations. However, neither the concentration of plasma free nor plasma total pentosidine at nadir correlated with serum creatinine. The intensity of immunostaining with anti-pentosidine antibody in proximal tubular cells was graded as weakly positive, positive, or strongly positive. Significant differences were obtained among plasma free pentosidine values between the weakly positive and strongly positive groups. CONCLUSIONS: Renal transplantation improves renal function and decreases renal excretion of free pentosidine. Accordingly, total pentosidine also decreases. However, the concentrations of plasma free and total pentosidine at nadir varied among individuals; the blood concentrations were not determined by renal function alone. It was suggested that deposition of pentosidine in proximal tubular cells was more severe among patients with higher plasma free pentosidine and serum creatinine values.