Effects of thyroxine on hyperkalemia and renal cortical Na+, K+ - ATPase activity induced by cyclosporin A

Cyclosporin A (CsA)-induced hyperkalemia is caused by alterations in transepithelial K+ secretion resulting from the inhibition of renal tubular Na+, K+ -ATPase activity. Thyroxine enhances renal cortical Na+, K+ -ATPase activity. This study investigated the effect of thyroxine on CsA-induced hyperkalemia. Sprague-Dawley rats were treated with either CsA, thyroxine, CsA and thyroxine, or olive-oil vehicle. CsA resulted in an increase in BUN and serum K+, along with a decrease in creatinine clearance, fractional excretion of potassium, and renal cortical Na+, K+ -ATPase activity, as compared with oil vehicle administration. Histochemical study showed reduced Na+, K+ -ATPase activity in the proximal tubular epithelial cells of the CsA-treated compared with the oil-treated rats. Histologically, isometric intracytoplasmic vacuolation, disruption of the arrangement and swelling of the mitochondria, and a large number of lysosomes in the tubular epithelium were characteristic of the CsA-treated rats. Co-administration of thyroxine prevented CsA-induced hyperkalemia and reduced creatinine clearance, Na+, K+ -ATPase activity, and severity of the histologic changes in the renal tubular cells when compared with the CsA-treated rats. Thyroxine increased the fractional excretion of potassium via the preservation of Na+, K+ -ATPase activity in the renal tubular cells. Thus, the beneficial effects of thyroxine may be suited to treatment modalities for CsA-induced hyperkalemia.     

MDM2 expression in EBV-infected nasopharyngeal carcinoma cells

To understand whether the p53-regulated mdm2 gene expression was altered by the Epstein-Barr virus (EBV) in nasopharyngeal carcinoma (NPC), the NPC-TW01 cell line was infected by EBV through IgA receptor-mediated endocytosis. The mdm2 gene was expressed only in a small fraction of the NPC cell population and could be enhanced in the EBV-infected (EBV+) cells. In the animals bearing EBV+ and EBV- NPC xenografts, the MDM2+ cells only appeared in clusters in both EBV+ and EBV- tumors with stronger expression in EBV+ cells. Cotransfection of pmdm2-Luc plus pSV40-p53 plus pCMV-LMP1 in the NPC-TW06 line that had p53 heterozygous point mutation showed stronger mdm2 promoter activity than cells cotransfected with pmdm2-Luc plus pSV40-p53, but no mdm2 promoter activity was seen in cells cotransfected with pmdm2-Luc plus pCMV-LMP1. Only the EBV-LMP1 but not the EBV-LMP2A gene could enhance p53 to upregulated mdm2 expression. Tumor cells in NPC biopsy specimens revealed similar mdm2 expression as in the animal model. It is concluded that although EBV can indirectly enhance mdm2 gene expression in tumor cells that express this gene, it cannot turn on or directly regulate mdm2 expression in cells that do not express this gene. In other words, EBV plays a role as an enhancer in NPC tumorigenesis.     

Hepatic and small bowel mucormycosis after chemotherapy in a patient with acute lymphocytic leukemia

Mucormycosis is a rare but invasive opportunistic fungal infection with increased frequency during chemotherapy-induced neutropenia. The clinical infections due to Mucor include rhinocerebral, pulmonary, cutaneous, gastrointestinal and disseminated diseases. The first two are the most common diseases and all entities are associated with a high mortality rate. Still hepatic involvement of Mucor is rarely reported. We experienced a case of hepatic and small bowel mucormycosis in a 56-year-old woman after induction chemotherapy for B-cell acute lymphocytic leukemia. Initial symptoms were a high fever unresponsive to broad spectrum antibiotics and pain in the left lower abdominal quadrant. It was followed by septic shock, deterioration of icterus and progressively elevated transaminase. An abdominal CT demonstrated multiple hypodense lesions with distinct margins in both lobes of liver and pericolic infiltration at small bowel and ascending colon. Diagnosis was confirmed by biopsy of the liver. The histopathology of the liver showed hyphae with the right-angle branching, typical of mucormycosis. The patient was managed with amphotericin B and operative correction of the perforated part of the small bowel was performed. However, the patient expired due to progressive hepatic failure despite corrective surgery and long-term amphotericin B therapy.     

Fatal pulmonary-renal syndrome manifested with immune complex crescentic glomerulonephritis in a patient with MPO-ANCA seropositivity

Recent reports have indicated that a significant number of immune complex glomerulonephritis (GN) cases are associated with antineutrophilic cytoplasmic antibody (ANCA). However, most of the reported cases were associated with underlying primary glomerular diseases. When primary glomerular diseases were not found, immune deposits tended to be non-specific and the level of ANCA is usually borderline. We report here upon a case of life-threatening pulmonary-renal syndrome manifested simultaneously with immune complex GN and myeloperoxidase (MPO)-ANCA seropositivity. A 29- year-old man was admitted with pulmonary hemorrhage and rapidly progressing renal dysfunction. On admission, ANCA revealed perinuclear staining with a titer of 1:160. The MPO-ANCA level was 59 IU by ELISA. Other serologic markers including ANA, anti-DS-DNA and anti-GBM Ab were negative. Renal biopsy showed cellular crescents in eight of 18 glomeruli. Immunofluorescence staining showed strong granular deposits of C3, C1q, IgG and IgM in the capillary loop and the mesangium. Electron microscopy showed multifocal electron dense deposits scattered in the mesangium, paramesangium, and the subendothelial and subepithelial areas. The patient initially responded to steroid and cyclophosphamide. MPO-ANCA decreased to less than 10 IU. Twenty three days after hospital discharge, the patient was re-admitted urgently with fever, generalized papulonodular skin lesions, and a recurrence of massive pulmonary hemorrhage and renal dysfunction. He died from uncontrolled pulmonary hemorrhage and respiratory insufficiency. P-ANCA titer and MPO-ANCA level at the second admission were 1:320 and 82 U/ml respectively. Interestingly, relapse was shown to be triggered by varicella zoster infection.     

Loss of tolerance to a maternal kidney transplant is selective for HLA class II: evidence from trans-vivo DTH and alloantibody analysis

We studied late graft rejection in a patient who had received a kidney transplant 9–10 years earlier from his mother and who had been off all immunosuppressive drugs for 7 years at the time of graft rejection onset. The mother differed for one HLA-A (A3) and one HLA-B (B62) antigen but had only a subtype mismatch at the HLA-DRβ1 locus (donor: DRβ1*1104; recipient: DRβ1*1102). A gradual rise in serum creatinine from 1.8 to 2.0 mg/dl at year 9 prompted a biopsy, which was negative for rejection (focal infiltrates but no tubulitis). Ten months later the patient’s creatinine had risen to > 3.4 mg/dl, and a second biopsy revealed extensive tubulitis, cellular rejection, and glomerular sclerosis. Sonicates of donor leukocytes triggered no delayed-type hypersensitivity (DTH) response above background (PBMC only) in the patient’s peripheral blood leukocytes obtained prior to year 9. A gradual recovery of antidonor DTH response between year 9 and 10 closely paralleled the change from tolerant to rejection status. Antidonor antibody was also undetectable in serum prior to year 9, but a donor-reactive antibody did develop at year 10.2 shortly after the peak of DTH response. The serum level of soluble donor HLA class I B62 antigen rose > 10-fold over prerejection level at the time of the biopsy-proven rejection, suggesting a possible trigger for both the cellular and humoral immune response. Nonetheless, we found no evidence for the de-velopment of humoral or cellular immunity to maternal HLA class I. Instead, DTH analysis of memory T cells of the patient obtained after rejection showed that a single maternal HLA DRβ1*1104 allopeptide, differing by two amino acids in sequence from the peptide of the recipient (DRβ1*1102), stimulated a strong memory DTH response. Similarly, we found an anti-HLA class II donor-specific antibody in serum that appeared to be crossreactive with DRβ1*1104 and DRβ1*1101 but not with the recipient DRβ1*1102 antigen. The data support the idea of a profound unresponsive state at both the cellular (DTH) and humoral level toward maternal HLA class I antigens that was not reversed even during late cellular rejection, despite the release of high levels of soluble HLA class I. Furthermore, the data suggest that DTH recovery was a close correlate of the onset of rejection and this “indirect” alloresponse, like the anti-donor alloantibody response that followed, was directed not to noninherited maternal HLA-A,B antigens but to the maternal HLA DRβ1*1104 subtype.     

Microdissection genotyping of archival fixative treated tissue for Gaucher disease

The genetic diagnosis of Gaucher disease by molecular methods is complicated by the existence of a highly homologous transcribed pseudogene (96% identity) that is found in close proximity to the true gene on chromosome 1q21. In addition, the pseudogene sequence can mimic disease-causing mutations in the true gene. Selective polymerase chain reaction (PCR) amplification of the true gene can be accomplished in extracted DNA from fresh-frozen samples by designing oligonucleotide primers to hybridize to defined regions that are not present in the pseudogene. This standard molecular approach, which entails amplification of relatively long segments of intact DNA, is not feasible in archival, paraffin-embedded, solid-tissue specimens in which the negative effects of chemical fixation result in DNA strand scission and breakdown of nucleic acid. A novel approach, specifically created for use with archival, fixative-treated tissue specimens, was developed for detection and characterization of common mutations of Gaucher disease. Three separate robust PCR reactions were formulated, 2 for selective amplification of portions of only the true gene exons 2 and 9, with a third reaction targeting exon 10, wherein both the true and pseudogene were coamplified. In the latter, DNA sequencing was used to determine the presence of true and pseudogene allele content in addition to identification of base sequence alterations. This method, requiring a single, 4-microm-thick histologic section, was successfully applied to archival paraffin block tissue specimens that had been in storage for up to 75 years. It was capable of accurately genotyping common Gaucher disease mutations as well as discovering a novel mutation and genetic polymorphism. We recommend our approach when only fixative-treated tis sue is available for molecular genotyping.     

Upregulation of VEGF and FGF2 in normal rat brain after experimental intraoperative radiation therapy

The expression of vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF)2 in the irradiated brain was examined to test how a single high dose radiation, similar to that used for intraoperative radiation therapy given to the normal cerebrum, can affect the vascular endothelium. After a burr hole trephination in the rat skull, the cerebral hemisphere was exposed to a single 10 Gy dose of gamma rays, and the radiation effect was assessed at 1, 2, 4, 6, and 8 weeks after irradiation. Histological changes, such as reactive gliosis, inflammation, vascular proliferation and necrosis, were correlated with the duration after irradiation. Significant VEGF and FGF2 expression in the 2- and 8-week were detected by enzyme-linked immunosorbent assay quantification in the radiation group. Immunohistochemical study for VEGF was done and the number of positive cells gradually increased over time, compared with the sham operation group. In conclusion, the radiation injuries consisted of radiation necrosis associated with the expression of VEGF and FGF2. These findings indicate that VEGF and FGF2 may play a role in the radiation injuries after intraoperative single high-dose irradiation.     

Expression of cell cycle regulators during smooth muscle cell proliferation after balloon catheter injury of rat artery

Intimal hyperplasia is defined as the abnormal migration and proliferation of vascular smooth muscle cells (VSMCs) with deposition of extracellular matrix. However, the cell cycle regulatory mechanisms of injury-induced VSMC proliferation are largely unknown. To examine the expression kinetics of cell cycle regulatory factors which is known to be worked positively or negatively, we used rat balloon injury model. Marked induction of proliferating cell nuclear antigen (PCNA), G1/S cyclin-dependent kinase (cdk2), and its regulatory subunit (cyclin E) occurred between 1 and 3 days after balloon arterial injury, and this was sustained for up to 7 days and then declined. However, the induction of the negative regulators, p21 and p27, occurred between 3 and 5 days of injury, peaked after 7 and 14 days and was then sustained. VSMC proliferation after balloon catheter injury of the rat iliac artery is associated with coordinated expression of positive (cdk2, cyclin E and PCNA) and negative (p21, p27) regulators. Cell cycle regulators such as cdk2, cyclin E, p21, p27 may be suitable targets for the control of intimal hyperplasia.     

APC I1307K and the E1317Q variants are not present in Chinese colorectal cancer patients.

PURPOSE: The APC I1307K and E1317Q variants predispose to colorectal adenomas and carcinomas in Caucasians, but data are lacking in Asians. METHODS AND RESULTS: We sequenced the APC gene from codons 1261 to 1409 and found none of 147 Chinese, 20 Malay, and 11 Indian colorectal cancer patients in Singapore to carry the APC I1307K or E1317Q variants. CONCLUSION: These variants are rare in these Asian populations, and play little role in colorectal cancer causation in Chinese.