Expression of urokinase-type plasminogen activator receptor (uPAR) in primary central nervous system neoplasms.

The cellular receptor for urokinase-type plasminogen activator receptor (uPAR) is a member of the glycosylphosphatidylinositol (GPI) anchored protein family. It is a specific cell surface receptor for its ligand, urokinase-type plasminogen activator, which catalyzes the formation of plasmin from plasminogen to generate the proteolytic cascade and leads to the breakdown of the extracellular matrix. uPAR has been shown to correlate with a propensity to tumor invasion and metastasis in several types of non-central nervous system tumors. In this study, the authors examined the immunohistochemical expression of uPAR in 65 primary brain tumors (5 pilocytic astrocytomas, 5 diffuse astrocytomas, 6 anaplastic astrocytomas, 8 glioblastomas, 5 oligodendrogliomas, 4 oligoastrocytomas, 6 anaplastic oligoastrocytomas, 4 gangliogliomas, 4 ependymomas, 5 medulloblastomas, 6 schwannomas, 5 meningiomas, 2 atypical meningiomas). The specimens were evaluated for intensity of immunostaining (0-3 scale), cellular localization of staining, and specific or unique patterns of staining. Some degree of uPAR expression was observed in all tumors. A significant positive correlation (P = 0.0006) between tumor grade and staining intensity was identified within the astrocytoma/glioblastoma subgroup, suggesting a possible correlation with anaplastic change and propensity to tumor invasion. Expression of uPAR in nonmalignant, noninvasive tumors such as schwannoma and meningioma suggests that uPAR may have other biologic functions in addition to promotion of tumor invasion.     

Effects of the fruit essential oil of Cuminum cyminum Linn. (Apiaceae) on acquisition and expression of morphine tolerance and dependence in mice

The problem of morphine tolerance and dependence is a universal phenomenon threatening social health everywhere the world. The major objective of this paper was to investigate the effects of fruit essential oil (FEO) of Cuminum cyminum on acquisition and expression of morphine tolerance and dependence in mice. Animals were rendered dependent on morphine using the well-established method in which was morphine (50, 50, 75 mg/kg; s.c.) injected three times daily for 3 days. In experimental groups, administration of FEO (0.001, 0.01, 0.1, 0.5, 1 and 2%; 5 ml/kg; i.p.) or Tween-80 (5 ml/kg; i.p.) was performed 60 min prior to each morphine injection (for acquisition) or the last injection of morphine on test day (for expression). Morphine tolerance was measured by tail-flick before and after administration of a single dose of morphine (50 mg/kg; s.c.) in test day (4th day). Morphine dependence was also evaluated by counting the number of jumps after injection of naloxone (5 mg/kg; i.p.) on the test day. The results showed that Cumin FEO, only at the dose of 2%, significantly attenuated the development of morphine tolerance (P < 0.01) and dependence (P < 0.05) while it could be significantly effective on expression of morphine tolerance (1 and 2%) and dependence (0.5, 1 and 2%) in a dose-dependent manner. Solely Cumin FEO injection (0.001–2%) did not show any analgesic effect. In conclusion, the essential oil of Cuminum cyminum seems to ameliorate the morphine tolerance and dependence in mice.     

A Strain Device Imposing Dynamic and Uniform Equi-Biaxial Strain to Cultured Cells

The objective of this study is to design a new apparatus to allow the control of the magnitude and frequency of dynamic stretch applied uniformly to cells cultured on a silicon elastic membrane. The apparatus is designed to produce equi-biaxial dynamic stretches with area changes ranging from 0% to 55% and frequencies ranging from 0 to 2 Hz. Homogeneous finite strain analysis using triangles of markers was performed to compute the symmetric two-dimensional Lagrangian strain tensor on the membrane. Measurements of strain in both static and dynamic conditions showed that the shear component of the strain tensor (E_rc) was near zero, and that there was no significant difference between radial (E_rr) and circumferential (E_cc) components, indicating the attainment of equi-biaxial strain. Bovine aortic endothelial cells were transiently transfected with a chimeric construct in which the luciferase reporter is driven by TPA-responsive elements (TRE). The transfected cells cultured on the membrane were stretched. The luciferase activity increased significantly only when the cells were stretched by 15% or more in area. Cells in different locations of the membrane showed similar induction of luciferase activities, confirming that strain is uniform and equi-biaxial across the membrane. © 1998 Biomedical Engineering Society. PAC98: 8780+s, 8745-k, 8722-q     

The Human MitoChip: a high-throughput sequencing microarray for mitochondrial mutation detection

Somatic mitochondrial mutations are common in human cancers, and can be used as a tool for early detection of cancer. We have developed a mitochondrial Custom Reseq microarray as an array-based sequencing platform for rapid and high-throughput analysis of mitochondrial DNA. The MitoChip contains oligonucleotide probes synthesized using standard photolithography and solid-phase synthesis, and is able to sequence >29 kb of double-stranded DNA in a single assay. Both strands of the entire human mitochondrial coding sequence (15,451 bp) are arrayed on the MitoChip; both strands of an additional 12,935 bp (84% of coding DNA) are arrayed in duplicate. We used 300 ng of genomic DNA to amplify the mitochondrial coding sequence in three overlapping long PCR fragments. We then sequenced >2 million base pairs of mitochondrial DNA, and successfully assigned base calls at 96.0% of nucleotide positions. Replicate experiments demonstrated >99.99% reproducibility. In matched fluid samples (urine and pancreatic juice, respectively) obtained from five patients with bladder cancer and four with pancreatic cancer, the MitoChip detected at least one cancer-associated mitochondrial mutation in six (66%) of nine samples. The MitoChip is a high-throughput sequencing tool for the reliable identification of mitochondrial DNA mutations from primary tumors in clinical samples.     

The role of COX-2 in angiogenesis and rheumatoid arthritis

Recent evidence suggests that cyclooxygenase (COX)-2 is a mediator of angiogenesis, and COX-2 activity is known to be upregulated in the rheumatoid arthritis (RA) synovium. We examined whether mediation of angiogenesis by COX-2 was occuring in cells of the RA synovium and in microvascular endothelial cells (ECs) that are similar to those found in the RA synovium. We demonstrate that rofecoxib, a selective COX-2 inhibitor, acts directly on human dermal microvascular ECs (HMVECs) to inhibit their chemotactic and tube forming ability. Likewise, pretreatment of HMVECs with rofecoxib significantly inhibited their ability to form tubes induced by conditioned media (CM) of activated RA synovial fibroblasts. When RA synovial fibroblasts were pretreated with rofecoxib for 16 h and then stimulated with interleukin (IL)-1beta, their CM induced significantly less HMVEC tube formation when compared with CM from vehicle-treated RA synovial fibroblasts. ELISAs performed on activated RA fibroblast CM for known proangiogenic factors demonstrated a significant reduction in bFGF, in addition to the expected decrease in PGE(2). Our studies suggest that COX-2-induced angiogenic activity is an active mechanism within diseased synovium and may provide an additional rationale for the use of COX-2 inhibitors in RA.     

Apparent genotype–phenotype correlation in childhood,adolescent, and adult Chediak‐Higashi syndrome

Chediak‐Higashi syndrome (CHS) is a rare autosomal recessive disorder characterized by severe immunologic defects, reduced pigmentation, bleeding tendency, and progressive neurological dysfunction. Most patients present in early childhood and die unless treated by bone marrow transplantation. About 10–15% of patients exhibit a much milder clinical phenotype and survive to adulthood, but develop progressive and often fatal neurological dysfunction. Very rare patients exhibit an intermediate adolescent CHS phenotype, presenting with severe infections in early childhood, but a milder course by adolescence, with no accelerated phase. Here, we describe the organization and genomic DNA sequence of the CHS1 gene and mutation analysis of 21 unrelated patients with the childhood, adolescent, and adult forms of CHS. In patients with severe childhood CHS, we found only functionally null mutant CHS1 alleles, whereas in patients with the adolescent and adult forms of CHS we also found missense mutant alleles that likely encode CHS1 polypeptides with partial function. Together, these results suggest an allelic genotype–phenotype relationship among the various clinical forms of CHS. © 2002 Wiley‐Liss, Inc.     

Genotype characterization and phylogenetic analysis of hepatitis B virus isolates from Iranian patients

Hepatitis B virus (HBV) is one of the major causative agents of acute and chronic liver disease worldwide and is believed to be responsible for a million deaths annually. Eight genotypes of HBV, A to H, have been described on the basis of similarity of the complete genomes sequence. Although, it is reported that the predominant HBV genotype in the Mediterranean area and the middle east is genotype D, there are no reports on HBV genotypes prevalent in Iran. In this study, the C and S regions of HBV from 26 chronic hepatitis B Iranian patients were amplified and sequenced. Phylogenetic analysis revealed that all Iranian HBV isolates sequences were classified into genotype D with bootstrap values of 100%, 73%, and 100% (1,000 replicates each) for S, C, and preS2 regions, respectively. The mean percent intra-distance of S and C regions were 0.8% and 2.3%, respectively. The mean percent inter-distance of S and C regions between Iranians and genotype D isolates were 1.7% and 3.0%, respectively, and the range of mean percent nucleotide distance of S and C regions between Iranians and the other reference isolates were 7.9%-17.5% and 4.8%-14.7%, respectively. Thirteen out of 23 HBV C region sequences showed nucleotide "A" at position 1896 (precore mutant) in C region. Nucleotide 1858 showed presence of "T" in all isolates. No insertion or deletion was found in both regions. SimPlot and BootScanning analyses did not show any recombination between Iranian isolates and other genotypes in both regions.     

Novel sequence variants in the TMC1 gene in Pakistani families with autosomal recessive hearing impairment

Though many hearing impairment genes have been identified, only a few of these genes have been screened in population studies. For this study, 168 Pakistani families with autosomal recessive hearing impairment not due to mutations in the GJB2 (Cx26) gene underwent a genome scan. Two-point and multipoint parametric linkage analyses were carried out. Twelve families had two-point or multipoint LOD scores of 1.4 or greater within the transmembrane cochlear expressed gene 1 (TMC1) region and were subjected to further screening with direct DNA sequencing. Five novel putatively functional non-synonymous sequence variants, c.830A>G (p.Y277C), c.1114G>A (p.V372M), c.1334G>A (p.R445H), c.2004T>G (p.S668R), and c.2035G>A (p.E679K), were found to segregate within seven families, but were not observed in 234 Pakistani control chromosomes. The variants c.830A>G (p.Y277C), c.1114G>A (p.V372M), and c.1334G>A (p.R445H) occurred at highly conserved regions and were predicted to lie within hydrophobic transmembrane domains, while non-synonymous variants c.2004T>G (p.S668R) and c.2035G>A (p.E679K) occurred in extracellular regions that were not highly conserved. There is evidence that the c.2004T>G (p.S668R) variant may have occurred at a phosphorylation site. One family has the known splice site mutation c.536 -8T>A. The prevalence of non-syndromic hearing impairment due to TMC1 in this Pakistani population is 4.4% (95%CI: 1.9, 8.6%). The TMC1 protein might have an important function in K(+) channels of inner hair cells, which would be consistent with the hypothetical structure of protein domains in which sequence variants were identified.     

Influence of age on iminodipropionitrile-induced vestibular and neurobehavioral toxicities in rats

A direct association between aging and drug-induced dyskinesia has been reported by several investigators. Iminiodipropionitrile (IDPN), a prototype nitrile compound produces a motor syndrome in rodents, which resembles neuroleptic drug induced dyskinesia. In this investigation attempt has been made to study the effect of age on IDPN induced vestibular hair cell degeneration and resulting dyskinetic syndrome. Male Wistar rats aged 3, 6 and 12 weeks received IDPN in the doses of 0, 200 and 400 mg/kg, intraperitoneally for 3 consecutive days. IDPN-induced dyskinesia was assessed using a behavioral testing battery on days 3, 4, 5, 6, 7, 14, 21 and 28. The rats were sacrificed on day 28; temporal bones were excised for vestibular histopathology and sera were collected for measuring the indices of oxidative stress (glutathione and conjugated dienes). IDPN in the dose of 200 mg/kg produced dyskinesia in 12 weeks old rats, but failed to do so in 3 and 6 weeks old rats. The high dose of IDPN (400 mg/kg) caused dyskinesia in all age groups, however, its onset and severity were age-dependent. Older rats showed an early onset and significantly high incidence of dyskinesia as compared to younger rats. The susceptibility of rats to IDPN-induced behavioral deficits was proportional to oxidative stress and degeneration of sensory hair cells in the crista ampullaris.