Biomedical subjects

Jun Zhang

Publications and source records attributed to Jun Zhang.

At least 73 records · Page 4Linked to original sources

PubMed2006

Reversible thermo-responsive sieving matrix for oligonucleotide separation.

A reversible thermo-responsive gel system, consisting of Pluronic copolymer mixture of F87 and F127, has been used to successfully carry out the separation of oligonucleotides, for the first time, by microchip-based capillary electrophoresis. Pluronic triblock copolymers F87 (E(61)P(40)E(61)) and F127 (E(99)P(69)E(99)), with E, P, and subscript denoting oxyethylene, oxypropylene, and segment length respectively, have a unique temperature dependent viscosity-adjustable property and a dynamic coating ability in aqueous solution, including 1 x TBE buffer. The mixture solution has a reversible thermo-responsive property and its sol-gel transition temperature can be adjusted ranging from about 17 degrees C to 38 degrees C by varying the relative weight ratio of F87 and F127 at an optimized concentration of approximately 30% (w/v) for oligonucleotide separations. Oligonucleotide sizing markers ranging from 8 to 32 base could be successfully separated in a 1.5 cm long separation channel by the mixture solution in its gel-like state. A 30% (w/v) with a F87/F127 weight ratio of 1 ratio 2 which has a "sol-gel" transition point of about 26 degrees C shows the best sieving ability. The sieving ability of the mixture solution was further confirmed in an Agilent Bioanalyzer 2100 system. Fast separation of oligonucleotides has been achieved within 40 s with one base resolution.

Electrophoresis, Microchip↗

PubMed2006

[The relationship between frameshift mutations of transforming growth factor-beta type II receptor, insulin growth factor II receptor, bcl-2 associated X protein, E2F4 and microsatellite instability in gastric carcinoma].

OBJECTIVE: To determine the microsatellite instability in gastric carcinomas, examine the frameshift mutations of transforming growth factor-beta type II receptor (TGFbetaRII), insulin growth factor II receptor (IGFIIR), bcl-2 associated X protein (BAX) and E2F4, and investigate whether or how alterations of the TGFbetaRII, IGFIIR, BAX and E2F4 gene are associated with MSI in gastric cancer. METHODS: Formalin-fixed, paraffin-embedded gastrectomy specimens and matching normal tissues of 65 cases of gastric carcinomas were retrieved from shanghai Ruijin Hospital and Shanghai East Hospital. DNA was extracted from tissue sections using phenol chloral isoamyl alcohol. Sections with no more than 50% of tumor cell areas were isolated by microdissection. DNA was amplified by PCR-based single strand conformation polymorphism (SSCP) for microsatellite analysis and was sequenced directly. Frameshift mutations in the coding regions, repetitive mononucleotide tracts of (A)10 for TGFbetaRII, (G)8 for IGFIIR, (G)8 for BAX, and trinucleotide repeats of (AGC)13 for transcription factors E2F4 were detected using PCR. Tumors were classified as being microsatellite stable (MSS) or having a low frequency of MSI (MSI-L, one of markers different in the tumor) or a high frequency of MSI (MSI-H, two or more of markers different). RESULTS: Eleven cases (18.0%) showed MSI-L, 12 (19.7%) showed MSI-H and 38 (62.3%) cases showed MSS. The mutation rates of TGFbetaRII, IGFIIR, BAX and E2F4 gene were 19.7%, 4.9%, 6.6% and 16.4% respectively. Among the 12 MSI-H gastric cancers, there were 10 TGFbetaRII mutations, 3 IGFIIR mutations, 4 BAX mutations and 10 E2F4 gene mutations. The alterations in the repeats of the related genes presented polymorphisms. Associations of MSI-H status and mutations of the 4 genes were highly significant (P < 0.01, respectively). No repeat tracts mutations in TGFbetaRII, IGFIIR, BAX and E2F4 gene were found in MSS tumors. CONCLUSIONS: The repeat coding regions within TGFbetaRII, IGFIIR, BAX and E2F4 gene are the targets of microsatellite instability. Frameshift mutations of the 4 genes play an important role in the development and progression of gastric cancers with microsatellite instability.

Adult↗

PubMed2006

CLIP-170 homologue and NUDE play overlapping roles in NUDF localization in Aspergillus nidulans.

Proteins in the cytoplasmic dynein pathway accumulate at the microtubule plus end, giving the appearance of comets when observed in live cells. The targeting mechanism for NUDF (LIS1/Pac1) of Aspergillus nidulans, a key component of the dynein pathway, has not been clear. Previous studies have demonstrated physical interactions of NUDF/LIS1/Pac1 with both NUDE/NUDEL/Ndl1 and CLIP-170/Bik1. Here, we have identified the A. nidulans CLIP-170 homologue, CLIPA. The clipA deletion did not cause an obvious nuclear distribution phenotype but affected cytoplasmic microtubules in an unexpected manner. Although more microtubules failed to undergo long-range growth toward the hyphal tip at 32 degrees C, those that reached the hyphal tip were less likely to undergo catastrophe. Thus, in addition to acting as a growth-promoting factor, CLIPA also promotes microtubule dynamics. In the absence of CLIPA, green fluorescent protein-labeled cytoplasmic dynein heavy chain, p150(Glued) dynactin, and NUDF were all seen as plus-end comets at 32 degrees C. However, under the same conditions, deletion of both clipA and nudE almost completely abolished NUDF comets, although nudE deletion itself did not cause a dramatic change in NUDF localization. Based on these results, we suggest that CLIPA and NUDE both recruit NUDF to the microtubule plus end. The plus-end localization of CLIPA itself seems to be regulated by different mechanisms under different physiological conditions. Although the KipA kinesin (Kip2/Tea2 homologue) did not affect plus-end localization of CLIPA at 32 degrees C, it was required for enhancing plus-end accumulation of CLIPA at an elevated temperature (42 degrees C).

Amino Acid Sequence↗

PubMed2006

The 2beta2-2beta3 loop of anthrax protective antigen contains a dominant neutralizing epitope.

Anthrax toxin consists of three proteins, protective antigen (PA), lethal factor, and edema factor. PA is the major component in the current anthrax vaccine, but the antigenic epitopes on it are not well-defined. We generated a pool of toxin-neutralizing anti-PA monoclonal antibodies (MAbs) to analyze the neutralizing epitopes of PA. Nine toxin-neutralizing MAbs obtained were found bound to three different domains of PA respectively, among which three MAbs with the strongest toxin-neutralizing activity recognized the same epitope within domain 2. This epitope was fine mapped to the chymotrypsin-sensitive site, (312)SFFD(315), in the 2beta(2)-2beta(3) loop of PA, using phage-displayed random peptide libraries and mutation analysis. The result demonstrated for the first time that the 2beta(2)-2beta(3) loop, which is involved in the transition of PA oligomers from prepore to pore, contains a dominant neutralizing epitope. This work contributes to the immunological and functional analysis of PA and offers perspective for the development of a new epitope vaccine against anthrax.

Animals↗

PubMed2006

Impact of cblB mutations on the function of ATP:cob(I)alamin adenosyltransferase in disorders of vitamin B12 metabolism.

ATP:cob(I)alamin adenosyltransferase (MMAB protein; methylmalonic aciduria type B) is an enzyme of vitamin B(12) metabolism that converts reduced cob(I)alamin to the adenosylcobalamin co-factor required for the functional activity of methylmalonyl-CoA mutase. Mutations in the human MMAB gene result in a block in adenosylcobalamin synthesis and are responsible for the cblB complementation group of inherited vitamin B(12) disorders. In this study, we examined the impact of several mutations, previously identified in cblB patients and clustered within a small, highly conserved region in MMAB. We confirmed mitochondrial expression of MMAB in human cells and showed that two mutations, R186W and E193K, were associated with absent protein by Western blot, while one, R191W, coupled with another point mutation, produced a protein in patient fibroblasts. Wild type MMAB and all four mutant proteins were stably expressed at high level as GST-fusion proteins, but only the R191W protein was enzymatically active. It showed an elevated K(m) of 320 microM (vs 6.8 microM for wild type enzyme) for ATP and 60 microM (vs 3.7 microM) for cob(I)alamin, with a reduction in k(cat) for both substrates. Circular dichroism spectroscopy revealed that three mutant proteins examined retained a alpha-helical structure as for the wild type protein. Characterization of MMAB will contribute to our understanding of cobalamin processing in mammalian cells and of disease mechanisms in the genetic disorders.

Alkyl and Aryl Transferases↗

PubMed2006

Oligomerization and aggregation of bovine pancreatic ribonuclease A: characteristic events observed by FTIR spectroscopy.

Nonnative protein aggregation, which is a common feature in biotechnology, is also a clinical feature in more than 20 serious degenerative diseases. We studied the specific events of bovine pancreatic ribonuclease A thermal aggregation by a combination of second derivative infrared analysis and two-dimensional infrared correlation spectroscopy. By comparing the events that occur in reversible and irreversible thermal unfolding processes, certain events that were related to protein aggregation were characterized. Particularly, a band that appeared at high temperatures was assigned to the cross beta-structures in oligomers. The effect of pH, NaCl, and ethanol on ribonuclease A oligomerization as well as further aggregation induced by heat were studied and dissimilar effects of these additives were found. Basic pH and NaCl could accelerate the thermal aggregation but did not affect the formation of oligomers, whereas ethanol could increase both the aggregation rate and the population of oligomers. Our results suggested that the aggregation of RNase A might be initiated by hydrophobic interactions, controlled by oligomerization and mediated by electrostatic interactions. Moreover, the strategy of using second derivative and two-dimensional infrared analysis might provide a potential powerful tool to study the events that are directly related to the initiation of protein aggregation.

Animals↗

PubMed2006

Down-regulation of hepatoma-derived growth factor inhibits anchorage-independent growth and invasion of non-small cell lung cancer cells.

We recently reported that a high level of hepatoma-derived growth factor (HDGF) expression in tumors correlates with a high incidence of tumor relapse or distant metastasis and shortened survival time in patients with non-small cell lung cancer (NSCLC). However, the mechanisms of the HDGF-associated aggressive biological behavior are unknown. In this study, we knocked down HDGF expression in NSCLC cells to determine the biological consequences. Transfection with HDGF-specific small interfering RNA (siRNA) resulted in down-regulation of HDGF expression in four NSCLC cell lines. Down-regulation of HDGF resulted in no detectable effect on anchorage-dependent cell growth as determined with a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, a microelectronic cell sensor system, and flow cytometry. In contrast, cells transfected with HDGF-siRNA grew more slowly and formed significantly fewer colonies in soft agar than did cells treated with LipofectAMINE alone or transfected with negative control siRNA. In an in vitro invasion assay, significantly fewer cells transfected with HDGF-siRNA than cells treated with LipofectAMINE alone were able to invade across a Matrigel membrane barrier. In an in vivo mouse model, A549 cells treated with HDGF-siRNA grown significantly slower than the cells treated with LipofectAMINE alone or negative control siRNA. Morphologically, HDGF-siRNA-treated tumors exhibited markedly reduced blood vessel formation and increased necrosis, whereas the Ki67 labeling indices were similar in tumors treated with controls. Our results suggest that HDGF is involved in anchorage-independent growth, cell invasion, and formation of neovasculature of NSCLC. These qualities may contribute to the HDGF-associated aggressive biological behavior of NSCLC.

Animals↗

PubMed2006

Nanostructured polymer matrix for oligonucleotide separation.

A nanostructured copolymer matrix has successfully separated oligonucleotides with high resolution by CE using a very short separation channel which simulates the real microchip condition for the first time. The triblock copolymer, E(45)B(14)E(45) (B20-5000) with E, B, and subscript denoting oxyethylene, oxybutylene, and segment length, respectively, has a unique temperature-dependent viscosity-adjustable property and a dynamic coating ability in 1xTBE buffer (89 mM Tris, 89 mM boric acid, 2 mM EDTA in Milli-Q water). The B-block is hydrophilic at low temperatures, e.g., 4 degrees C, and the polymer solution has a very low viscosity of about 100 cP in a 32.5% w/v solution. At room temperatures, the B-block becomes hydrophobic due to a breakdown of hydrogen bonds between B-blocks and water, and the polymer matrix forms a body-centered cubic structure at high concentrations. Oligonucleotide sizing markers ranging from 8 to 32 base could be successfully separated with one-base resolution in a 1.5 cm long separation channel by E(45)B(14)E(45) in its gel-like state.

Electrophoresis, Capillary↗

PubMed2006

Scale-up development of high-performance polymer matrix for DNA sequencing analysis.

Linear polyacrylamide (LPA) has been widely used as a replaceable separation matrix in CE. An increase in the molecular weight of the separation medium favors the separation of larger DNA fragments. In order to obtain ultrahigh-molecular-weight (UHMW) LPA, a "frozen" method was developed to synthesize the LPA homopolymer. This approach has three major advantages when compared with other existing routes of LPA synthesis: (i) long LPA chains could be obtained easily, with their average molecular weight (MW) being in the high 10 MDa range; (ii) the desired MW could be adjusted over a broad range by controlling the temperature and the concentration of initiators during synthesis; (iii) the product solution contains only a tiny amount of impurity besides the solvent and LPA. Both static and dynamic laser light scattering measurements were carried out to characterize the synthesized LPA in the buffer solution. The DNA sequencing matrix prepared from LPA using this method was studied and the results were compared with the newly developed commercial product POP7 from Applied Biosystems. It should be noted that this approach can be applied to synthesize other water-soluble polymers, resulting in UHMW products because the chain transfer constant is smaller at lower temperatures.

Acrylic Resins↗

PubMed2006

A new inducible adenoviral expression system that responds to inflammatory stimuli in vivo.

BACKGROUND: Gene transfer using inducible promoters, which control expression of transgenic proteins in response to physiological conditions, may have significant advantages. In this study, we tried to achieve an inducible adenoviral expression system for physiologically responsive gene therapy of autoimmune or inflammatory diseases. METHODS: A luciferase reporter vector with a hybrid promoter containing the human IL-1beta enhancer region (-3690 to - 2720) and the human CIITA promoter IV (-399 to + 2) was constructed. A replication-deficient adenovirus was engineered with luciferase controlled by the IL1beta/CIITApIV promoter (Ad-IL1beta/CIITApIV-Luc). The reporter vector or adenovirus was transfected to C57Bl/6 myeloid dendritic cells (DCs), RAW264.7, and Hep G2 to study the in vitro characteristics of this hybrid promoter. An inflammation model was prepared by injecting lipopolysaccharide (LPS) into Balb/c mice intraperitoneally (i.p.), and infected with Ad-IL1beta/CIITApIV-Luc or Ad-CMV-Luc to study the in vivo characteristics of the IL1beta/CIITApIV promoter. RESULTS: The IL1beta/CIITApIV hybrid promoter has pronounced promoter activity, broad-range responsiveness to cytokines or LPS, and can be rechallenged after first induction. In the inflammation model, IL1beta/CIITApIV could drive hepatic luciferase expression increasedly rapidly after LPS challenge and in a LPS dose-dependent manner. CONCLUSIONS: Using the IL1beta/CIITApIV hybrid promoter in gene transfer vectors may make it possible to produce transgenic proteins in vivo in direct relationship with the intensity and duration of an individual's status. By providing endogenously controlled production of transgenic proteins, this approach might limit the severity of autoimmune or inflammatory response without interfering with the beneficial components of host defense and immunity.

Adenoviridae↗

PubMed2006

Determination of characteristics of degenerative joint disease using optical coherence tomography and polarization sensitive optical coherence tomography.

BACKGROUND AND OBJECTIVES: Previous studies have demonstrated that optical coherence tomography (OCT) could be used to delineate alterations in the microstructure of cartilage, and have suggested that changes in the polarization state of light as detected by OCT could provide information on the birefringence properties of articular cartilage as influenced by disease. In this study we have used both OCT and polarization sensitive optical coherence tomography (PS-OCT) technologies to evaluate normal and abnormal bovine articular cartilage according to established structural, organizational, and birefringent characteristics of degenerative joint disease (DJD) in order to determine if this technology can be used to differentiate various stages of DJD as a minimally invasive imaging tool. MATERIALS AND METHODS: Fresh bovine femoral-tibial joints were obtained from an abattoir, and 45 cartilage specimens were harvested from 8 tibial plateaus. Whole ex vivo specimens of normal and degenerative articular cartilage were imaged by both OCT and PS-OCT, then fixed and processed for histological evaluation. OCT/PS-OCT images and corresponding histology sections of each specimen were scored according to a modified Mankin structural grading scale and compared. RESULTS: OCT and PS-OCT imaging allowed structural evaluation of intact articular cartilage along a 6 mm surface length to a depth of 2 mm with a transverse resolution of 12 microm and an axial resolution of 10 microm. The OCT and PS-OCT images demonstrated characteristic alterations in the structure of articular cartilage with a high correlation to histological evaluation (kappa = 0.776). The OCT images were able to demonstrate early to advanced structural changes of articular cartilage while the optical phase retardation images obtained by PS-OCT imaging were able to discriminate areas where disorganization of the cartilage matrix was present, however, these characteristics are much different than those reported where OCT images alone were used to characterize tissue birefringence. No evidence of differences in OCT or PS-OCT images were detected between specimens of similar structural characteristics where proteoglycan was judged present or absent by safranin-O Fast Green staining. CONCLUSIONS: The combined use of OCT and PS-OCT technologies to obtain images from a single system is able to demonstrate and discriminate between characteristics of very early stages of surface irregularities not previously reported for OCT imaging, to deep clefts and collagen matrix disorganization for tissue at depths of up to 2 mm with good correlation to histology. PS-OCT and accumulated optical phase retardation images of articular cartilage as constructed from alterations in Stokes vector parameters appear to give a valuable but different assessment of alterations in tissue birefringence and organization than have been reported for OCT images obtained with the use of polarized or non-polarized light sources. This is the first time that alterations in the polarization state of light reflected from within the tissue have been demonstrated to be consistent with changes observed in the orientation and organization of the collagen matrix in advanced stages of DJD. The degree of phase transformation of light reflected from within the tissue as determined by PS-OCT imaging does not appear to be altered by the presence or absence of proteoglycan.

Animals↗

PubMed2006

A high-throughput liquid chromatography/tandem mass spectrometry method for simultaneous quantification of a hydrophobic drug candidate and its hydrophilic metabolite in human urine with a fully automated liquid/liquid extraction.

ABT-869 (A-741439) is an investigational new drug candidate under development by Abbott Laboratories. ABT-869 is hydrophobic, but is oxidized in the body to A-849529, a hydrophilic metabolite that includes both carboxyl and amino groups. Poor solubility of ABT-869 in aqueous matrix causes simultaneous analysis of both ABT-869 and its metabolite within the same extraction and injection to be extremely difficult in human urine. In this paper, a high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS) method has been developed and validated for high-speed simultaneous quantitation of the hydrophobic ABT-869 and its hydrophilic metabolite, A-849529, in human urine. The deuterated internal standards, A-741439D(4) and A-849529D(4), were used in this method. The disparate properties of the two analytes were mediated by treating samples with acetonitrile, adjusting pH with an extraction buffer, and optimizing the extraction solvent and mobile phase composition. For a 100 microL urine sample volume, the lower limit of quantitation was approximately 1 ng/mL for both ABT-869 and A-849529. The calibration curve was linear from 1.09 to 595.13 ng/mL for ABT-869, and 1.10 to 600.48 ng/mL for A-849529 (r2 > 0.9975 for both ABT-869 and A-849529). Because the method employs simultaneous quantification, high throughput is achieved despite the presence of both a hydrophobic analyte and its hydrophilic metabolite in human urine.

Chromatography, High Pressure Liquid↗

PubMed2006

Method development and validation for zotarolimus concentration determination in stented swine arteries by liquid chromatography/tandem mass spectrometry detection.

Drug-eluting stents have attracted significant attention in the medical community and pharmaceutical industry due to their proven success in significantly reducing restenosis. Abbott Laboratories is developing a drug-eluting stent coated with zotarolimus and swine was recently used as an animal model for the pre-clinical study of stent implantation. In this article, we present a detailed experimental design and results for the validation and sample analysis of zotarolimus drug concentration in stented swine artery samples. Introduction of tissue quality control (QC) samples allows evaluation of the entire analytical process as well as the stability of the drug in both original tissue and homogenized tissue samples. In addition, a novel approach using 100% swine blood as the homogenization solution was developed for the consistency of the liquid-liquid extraction recovery and stability of the zotarolimus in tissue homogenates. Standards were prepared by spiking zotarolimus working solution in swine blood and tissue QC samples were used along with the artery samples during the sample analysis. The linear dynamic range of blood standard samples is from 0.61 to 333.20 ng/mL to accommodate the predicted artery homogenate concentrations. Overall tissue QC %CV during the method validation was from 4.4% to 8.6%. The overall %bias of tissue QC samples during the method validation was from -7.3% to 16.6%. The method was successfully applied for the analysis of swine artery samples. A similar approach for method validation and sample analysis has been successfully applied for the analysis of swine myocardium, kidney and liver tissue samples.

Animals↗

PubMed2006

Developing multiple high-throughput GLP methods for an investigational drug candidate in various matrices within a 96-well plate.

In early pharmaceutical product development, an investigational drug candidate is typically dosed to various species for toxicological and pharmacokinetic studies. Most of these studies require multiple analytical methods that have to be validated with good laboratory practice (GLP) prior to the application in regulated studies. Usually, these analytical methods are developed in either a serial or parallel approach. For either approach, the development of multiple analytical methods takes tremendous work from scientists and instruments, and thus is not cost-effective. In this respect, a new strategy has been developed for simultaneous GLP method development using liquid chromatographic separation and tandem mass spectrometric detection. This high-throughput approach allows system suitability, carryover, calibration curve, accuracy, precision, matrix effect and selectivity to be evaluated in one 96-well plate. The strategy has been successfully implemented for multiple investigational drug candidates at Abbott Laboratories. The methods developed with this strategy are accurate, precise, selective, robust and matrix-independent. As an example, ABT-279 was used to demonstrate the feasibility of this strategy.

Chromatography, Liquid↗

PubMed2006

Retrospective comparison of retroperitoneal laparoscopic versus open dismembered pyeloplasty for ureteropelvic junction obstruction.

PURPOSE: We evaluated the clinical value of retroperitoneal laparoscopic dismembered pyeloplasty for ureteropelvic junction obstruction compared with open surgery. MATERIALS AND METHODS: The clinical data of 56 patients who underwent retroperitoneal laparoscopic dismembered pyeloplasty were retrospectively compared with those of 40 patients who underwent open dismembered pyeloplasty through a retroperitoneal flank approach. The Student t test, Pearson chi-square test and Mann-Whitney rank sum test were applied for statistical analysis as appropriate. RESULTS: Patient demographic data were similar between the 2 groups. In the laparoscopic group operative time (80 vs 120 minutes), estimated blood loss (10 vs 150 ml), recovery of intestinal function (1 vs 2 days), analgesic requirements (diclofenac sodium suppository) (75 vs 150 mg), incision length (3.5 vs 21 cm) and postoperative hospital stay (7 vs 9 days) were better than in the open group (p <0.001 for all). No intraoperative complications occurred in either group. The incidence of postoperative complications (2 of 56, 3.6% vs 3 of 40, 7.5%, p = 0.729) and success rates (55 of 56, 98.2% vs 39 of 40, 97.5%, p = 0.058) were equivalent in the 2 groups. CONCLUSIONS: Retroperitoneal laparoscopic dismembered pyeloplasty is a minimally invasive, safe and effective therapy for ureteropelvic junction obstruction with low morbidity, shorter convalescence and excellent outcomes, and can be accomplished reasonably quickly in experienced hands.

Adult↗

PubMed2006

Dynamic contrast enhanced magnetic resonance imaging as a biological marker to noninvasively assess the effect of finasteride on prostatic suburethral microcirculation.

PURPOSE: We assessed dynamic contrast enhanced magnetic resonance imaging as a biological marker of in vivo changes in microcirculation in the prostatic suburethral region. MATERIALS AND METHODS: A total of 12 male beagle dogs with spontaneous benign prostatic hyperplasia were randomly allocated to 1 control group and 1 finasteride (Merck and Co., Whitehouse Station, New Jersey) treated group. Two baseline dynamic contrast enhanced magnetic resonance imaging examinations and 3 followups were performed to assess prostate microcirculation. Treatment duration was 3 months. The pharmacokinetic parameters evaluated in prostatic suburethral areas were the maximum enhancement ratio in AU, time to maximum signal enhancement in minutes, amplitude in AU and the exchange rate constant in minutes(-1). RESULTS: After completion of the therapeutic regimen time to maximum signal enhancement was significantly longer in the finasteride group than in controls (p < 0.01). Amplitude and the exchange rate constant decreased 39% and 34%, respectively, in the finasteride group at the end of treatment, which significantly differed from results in the control group (p < 0.05). CONCLUSIONS: Dynamic contrast enhanced magnetic resonance imaging is capable of noninvasively assessing the prostatic microcirculation changes induced by finasteride. Pharmacokinetic parameters show considerable promise to be biomarkers for the development of benign prostatic hyperplasia drugs such as 5alpha-reductase inhibitors by the in vivo monitoring of microvascular changes. A relevant clinical application could be the pretreatment assessment of finasteride effectiveness to decrease perioperative bleeding at transurethral prostate resection and in treatment for hematuria.

Animals↗

PubMed2006

A live cell, image-based approach to understanding the enzymology and pharmacology of 2-bromopalmitate and palmitoylation.

The addition of a lipid moiety to a protein increases its hydrophobicity and subsequently its attraction to lipophilic environments like membranes. Indeed most lipid-modified proteins are localized to membranes where they associate with multiprotein signaling complexes. Acylation and prenylation are the two common categories of lipidation. The enzymology and pharmacology of prenylation are well understood but relatively very little is known about palmitoylation, the most common form of acylation. One distinguishing characteristic of palmitoylation is that it is a dynamic modification. To understand more about how palmitoylation is regulated, we fused palmitoylation substrates to fluorescent proteins and reported their subcellular distribution and trafficking. We used automated high-throughput fluorescence microscopy and a specialized computer algorithm to image and measure the fraction of palmitoylation reporter on the plasma membrane versus the cytoplasm. Using this system we determined the residence half-life of palmitate on the dipalmitoyl substrate peptide from GAP43 as well as the EC(50) for 2-bromopalmitate, a common inhibitor of palmitoylation.

Amino Acid Motifs↗

PubMed2006

Mechanisms and biomarkers of cardiovascular injury induced by phosphodiesterase inhibitor III SK&F 95654 in the spontaneously hypertensive rat.

The cardiovascular injury of the type III selective PDE inhibitor SK&F 95654 was investigated in SHR. Twenty-four hours after a single sc injection of 100 or 200 mg/kg of the drug, rats exhibited cardiomyocyte necrosis and apoptosis, interstitial inflammation, hemorrhage and edema, as well as mesenteric arterial hemorrhage and necrosis, periarteritis, EC and VSMC apoptosis, EC activation, and MC activation and degranulation. Elevated serum levels of cTnT and decreased cTnT immunoperoxidase staining on cardiomyocytes were detected in the drug-treated rats. Serum levels of alpha2-macroglobulin and IL-6 were significantly elevated following drug treatment. NMR spectral patterns of urine samples are significantly different between the drug-treated and control rats. These results indicate that measurement of serum cTnT, acute phase proteins, and cytokines as well as metabonomic urine profiles may serve as potential biomarkers for drug-induced cardiovascular injury in rats. Increased expression of CD63 on MC (tissue biomarker of MC), of nitrotyrosine on MC and EC (an indirect indicator of NO in vivo), and of iNOS on MC and EC (source of NO) suggest that NO produced by activated and degranulated MC as well as activated EC play an important role in SK&F 95654-induced mesenteric vascular injury.

Alanine Transaminase↗