Biomedical subjects

G Q Wang

Publications and source records attributed to G Q Wang.

At least 19 recordsLinked to original sources

PubMed2006

Role of cytosolic liver fatty acid binding protein in hepatocellular oxidative stress: effect of dexamethasone and clofibrate treatment.

The presence of cysteine and methionine groups together with an ability to bind long-chain fatty acid (LCFA) oxidation products makes liver fatty acid binding protein (L-FABP) an attractive candidate against hepatocellular oxidative stress. In this report, we show that pharmacological treatment directed at modulating L-FABP level affected hepatocellular oxidant status. L-FABP expressing 1548-hepatoma cells, treated with dexamethasone or clofibrate, decreased and increased intracellular L-FABP levels, respectively. Oxidative stress was induced by H2O2 incubation or hypoxia-reoxygenation. The fluorescent marker, dichlorofluorescein (DCF), was employed to measure intracellular reactive oxygen species (ROS). Hepatocellular damage was assessed by lactate dehydrogenase (LDH) level. Dexamethasone treatment resulted in a significant increase in DCF fluorescence with higher LDH release compared to control cells. Clofibrate treatment, however, resulted in a significant decrease in both parameters (p<0.05). Drug treatments did not affect cytosolic activities of glutathione peroxidase (GPx), superoxide dismutase (SOD), or catalase suggesting that the differences between treated and control cells may likely be associated with varying L-FABP levels. We conclude that L-FABP may act as an effective endogenous cytoprotectant against hepatocellular oxidative stress.

Carcinoma, Hepatocellular↗

PubMed2006

An integrated numerical and physical modeling system for an enhanced in situ bioremediation process.

Groundwater contamination due to releases of petroleum products is a major environmental concern in many urban districts and industrial zones. Over the past years, a few studies were undertaken to address in situ bioremediation processes coupled with contaminant transport in two- or three-dimensional domains. However, they were concentrated on natural attenuation processes for petroleum contaminants or enhanced in situ bioremediation processes in laboratory columns. In this study, an integrated numerical and physical modeling system is developed for simulating an enhanced in situ biodegradation (EISB) process coupled with three-dimensional multiphase multicomponent flow and transport simulation in a multi-dimensional pilot-scale physical model. The designed pilot-scale physical model is effective in tackling natural attenuation and EISB processes for site remediation. The simulation results demonstrate that the developed system is effective in modeling the EISB process, and can thus be used for investigating the effects of various uncertainties.

Biodegradation, Environmental↗

PubMed2005

Membrane binding proteins are the major determinants for the hepatocellular transmembrane flux of long-chain fatty acids bound to albumin.

PURPOSE: The hepatic transmembrane flux of long-chain fatty acids (LCFA) occurs through passive and fatty acid transport protein facilitated processes from blood. The extent that these transport processes can be related to the unbound and protein-bound fractions of LCFA in blood is not clear. METHODS: We used hepatocyte suspensions, hepatoma monolayers, and perfused rat livers to quantitate the transport of purified [(3)H]palmitate ([(3)H]PA) and 12-(N-methyl)-N-[(7-nitrobenz-2oxa-1,3-diazol-4yl-)amino]octadecanoicacid (12-NBDS) from solutions with a constant unbound LCFA concentration with varying bovine serum albumin (BSA) concentrations and in the presence and absence of antisera raised against cytosolic liver fatty acid binding protein (L-FABP). RESULTS: In the absence of L-FABP antisera, using an unbound ligand concentration that was adjusted to remain constant at each BSA concentration, hepatocyte [(3)H]PA and 12-NBDS uptake rates increased linearly with an increase in BSA concentration (p < 0.0001). In the presence of L-FABP antisera, [(3)H]PA uptake showed a greater reduction in the presence of 100 muM BSA than 5 muM BSA. The calculated permeability surface area product (PS) confirmed that both unbound and bound fractions of LCFA contributed to the overall flux, but only the PS for the protein-bound fraction was reduced in the presence of L-FABP antisera. In situ rat liver perfusion studies showed that the only rate process for the disposition of [(3)H]PA in the liver inhibited by L-FABP antisera was that for influx, as defined by PS, and that it reduced PS in the perfused liver by 42%. CONCLUSION: These results suggest that, at physiological albumin concentrations, most of the LCFA uptake is mediated from that bound to albumin by a hepatocyte basolateral membrane transport protein, and uptake of unbound LCFA occurring by passive diffusion contributes a minor component.

Albumins↗

PubMed2005

Assessing the cellular transmembrane electrical potential difference on the hepatic uptake of palmitate.

Understanding the driving forces for the hepatic uptake of endogenous and exogenous substrates in isolated cells and organs is fundamental to describing the underlying hepatic physiology/pharmacology. In this study we investigated whether uptake of plasma protein-bound [3H]-palmitate across the hepatocyte wall is governed by the transmembrane electrical potential difference (PD). Uptake was studied in isolated hepatocytes and isolated perfused rat livers (IPL). Protein-binding and vasoactive properties of the different perfusates were determined using in vitro heptane/buffer partitioning studies and the multiple indicator dilution (MID) technique in the IPL, respectively. Altering hepatocyte PD by perfusate ion substitution resulted in either a substantial depolarization (-14 +/- 1 mV, n = 12, mean +/- S.E., substituting choline for Na+) or hyperpolarization (-46 +/- 3 mV, n = 12, mean +/- S.E., substituting nitrate for Cl-). Perfusate ion substitution also affected the equilibrium binding constant for the palmitate-albumin complex. IPL studies suggested that, other than with gluconate buffer, hepatic [3H]-palmitate extraction was not affected by the buffer used, implying PD was not a determinant of extraction. [3H]-Palmitate extraction was much lower (p < 0.05) when gluconate was substituted for Cl- ion. This work contrasts with that for the extraction of [3H]-alanine where hepatic extraction fraction was significantly reduced during depolarization. Changing the albumin concentration did not affect hepatocyte PD, and [3H]-palmitate clearance into isolated hepatocytes was not affected by the buffers used. MID studies with vascular and extravascular references revealed that, with the gluconate substituted buffer, the extravascular volume possibly increased the diffusional path length thus explaining reduced [3H]-palmitate extraction fraction in the IPL.

Alanine↗

PubMed2003

Cladistic analysis of human apolipoprotein a4 polymorphisms in relation to quantitative plasma lipid risk factors of coronary heart disease.

Genetic variation in several genes involved in lipid metabolism is known to affect population variation in quantitative lipid risk factor profiles for coronary heart disease (CHD). The apolipoprotein A-IV gene (APOA4) is one such candidate gene. We genotyped five polymorphisms in the APOA4 gene (codon 127, codon 130, codon347, codon 360 and 3' VNTR) and investigated their impact on plasma lipid trait levels in three populations comprising 604 U.S. non-Hispanic Whites (NHWs), 408 U.S. Hispanics and 708 Nigerian Blacks. Cladistic analysis was carried out to identify 5-site haplotypes that were associated with significant phenotypic differences in each population. The distribution of APOA4 genotypes was significantly different between ethnic groups. The Africans were monomorphic for two of the five sites (codons 130 and 360), but possess a unique 12 bp insertion that was not observed in NHWs and Hispanics. Due to linkage disequilibrium between the sites, only 6 haplotypes were observed in NHWs and Hispanics, and 4 in Africans. Several gender-and ethnic-specific associations between genotypes and plasma lipid traits were observed when single sites were used. Several haplotypes were identified by cladistic analysis that may carry functional mutations that affect plasma lipid trait levels.

Adult↗

PubMed2001

Resistance to granzyme B-mediated cytochrome c release in Bak-deficient cells.

Granzyme B (GrB), a serine protease with substrate specificity similar to the caspase family, is a major component of granule-mediated cytotoxicity of T lymphocytes. Although GrB can directly activate caspases, it induces apoptosis predominantly via Bid cleavage, mitochondrial outer membrane permeabilization, and cytochrome c release. To study the molecular regulators for GrB-mediated mitochondrial apoptotic events, we used a CTL-free cytotoxicity system, wherein target cells are treated with purified GrB and replication-deficient adenovirus (Ad). We report here that the Bcl-2 proapoptotic family member, Bak, plays a dominant role in GrB-mediated mitochondrial apoptotic events. A variant of Jurkat cells, deficient in Bak expression, was resistant to GrB/Ad-mediated apoptosis, as determined by lack of membranous phosphatidylserine exposure, lack of DNA breaks, lack of mitochondrial outer membrane permeabilization, and unchanged expression of inner mitochondrial membrane cardiolipin. The resistance of Bak-deficient cells to GrB/Ad cytotoxicity was reversed by transduction of the Bak gene into these cells. The requirement for both Bid and Bak, was further demonstrated in a cell-free system using purified mitochondria and S-100 cytosol. Purified mitochondria from Bid knockout mice, but not from Bax knockout mice, failed to release cytochrome c in response to autologous S-100 and GrB. Also, Bak-deficient mitochondria did not release cytochrome c in response to GrB-treated cytosol unless recombinant Bak protein was added. These results are the first to report a role for Bak in GrB-mediated mitochondrial apoptosis. This study demonstrates that GrB-cleaved Bid, which differs in size and site of cleavage from caspase-8-cleaved Bid, utilizes Bak for cytochrome c release, and therefore, suggests that deficiency in Bak may serve as a mechanism of immune evasion for tumor or viral infected cells.

Adenoviridae↗

PubMed2001

A role for mitochondrial Bak in apoptotic response to anticancer drugs.

In the present study a clonal Jurkat cell line deficient in expression of Bak was used to analyze the role of Bak in cytochrome c release from mitochondria. The Bak-deficient T leukemic cells were resistant to apoptosis induced by UV, staurosporin, VP-16, bleomycin, or cisplatin. In contrast to wild type Jurkat cells, these Bak-deficient cells did not respond to UV or treatment with these anticancer drugs by membranous phosphatidylserine exposure, DNA breaks, activation of caspases, or release of mitochondrial cytochrome c. The block in the apoptotic cascade was in the mitochondrial mechanism for cytochrome c release because purified mitochondria from Bak-deficient cells failed to release cytochrome c or apoptosis-inducing factor in response to recombinant Bax or truncated Bid. The resistance of Bak-deficient cells to VP-16 was reversed by transduction of the Bak gene into these cells. Also, the cytochrome c releasing capability of the Bak-deficient mitochondria was restored by insertion of recombinant Bak protein into purified mitochondria. Following mitochondrial localization, low dose recombinant Bak restored the mitochondrial release of cytochrome c in response to Bax; at increased doses it induced cytochrome c release itself. The function of Bak is independent of Bid and Bax because recombinant Bak induced cytochrome c release from mitochondria purified from Bax(-/-), Bid(-/-), or Bid(-/-) Bax(-/-) mice. Together, our findings suggest that Bak plays a key role in the apoptotic machinery of cytochrome c release and thus in the chemoresistance of human T leukemic cells.

Adenoviridae↗

PubMed2001

Cardiac apoptosis in burned rats with delayed fluid resuscitation.

Clinical and experimental studies have shown that delayed fluid resuscitation postburn decreases heart function. We hypothesized that apoptosis occurs in the cardiomyocyte in this condition. To investigate this hypothesis, rats were burned, fluid resuscitation was delayed, and the integrity of cardiac genomic DNA in the burned rats was determined with an LM-PCR Ladder Assay kit. DNA fragmentation shown as DNA ladders on gels, the hallmark of apoptosis, was found in the heart tissue of these rats. In the early phase of delayed fluid resuscitation, the nuclear factor kappa B (NF-kappa B) was examined using an electrophoretic mobility shift assay and was found to be activated. In comparison with burned rats with immediate fluid resuscitation, nitric oxide levels in hearts from burned rats with delayed fluid resuscitation were significantly lower (P<0.01). These results suggest that apoptosis may be an important pathway for cardiac injury, which may result from the activation of NF-kappa B and decreased nitric oxide levels.

Animals↗

PubMed2001

Palmitate uptake by neonatal rat myocytes and hepatocytes. Role of extracellular protein.

The role of extracellular binding proteins in the rate of [3H]palmitate uptake by neonatal cardiac myocytes and hepatocytes was investigated using a model-independent approach. Binding proteins used in this study included alpha1-acid glycoprotein [isoelectric point (pI) approximately 2.7], conalbumin (pI approximately 6.4), lysozyme (pI approximately 11.0), albumin (pI approximately 4.9), and albumin which had been modified to yield proteins with pI values of 3.5, 4.7, 7.5 and 8.6. All uptake studies were conducted at similar unbound ligand fractions. There was a linear relationship between the rate of neonatal hepatocyte [3H]palmitate clearance and protein pI (r2 = 0.98). In contrast, there was an overall poor relationship between neonatal cardiac myocyte [3H]palmitate-clearance rate and protein pI (r2 = 0.48). However, the relationship improved when the data on [3H]palmitate-clearance were analyzed using only the modified albumins. The study indicates that an ionic interaction between extracellular proteins and the hepatocyte surface enhances the overall uptake of [3H]palmitate. This interaction may be limited to albumin for neonatal cardiac myocytes.

Animals↗

PubMed2001

Tumor necrosis factor-alpha-promoted expression of Bcl-2 and inhibition of mitochondrial cytochrome c release mediate resistance of mature dendritic cells to melanoma-induced apoptosis.

Melanoma escapes host defenses through a variety of means, including the elimination of immune effector cells within the tumor microenvironment. We have reported recently that murine and human tumors including melanoma induce premature apoptosis of dendritic cells both in vitro and in vivo. In this study, we have demonstrated that overexpression of the Bcl-2 protein family member Bcl-xL rescued murine dendritic cells (DCs) from melanoma-induced death in vitro. Another successful protection approach was tumor necrosis factor (TNF)-alpha-promoted sustained expression of the antiapoptotic protein Bcl-2 within dendritic cells. This effect of TNF-alpha was mediated by inhibition of mitochondrial cytochrome c release. Thus, both Bcl-xL and Bcl-2 enhance survival of dendritic cells within the tumor microenvironment. In addition, mature DCs were more resistant to melanoma-induced apoptosis than immature dendritic cells. This finding suggests a stage-dependent sensitivity of DCs to tumor-induced cell death. We conclude that: (a) mature DCs might be more suitable for the use of cancer vaccination; and (b) Bcl-2 protein family members such as Bcl-xL and Bcl-2 rescue DCs from tumor-induced premature apoptosis.

Animals↗

PubMed2001

Hepatocyte [3H]-palmitate uptake: effect of albumin surface charge modification.

The role of plasma proteins on the cellular uptake of lipophilic substrates has perplexed investigators for many years. We tested the hypothesis that an ionic interaction between the protein-ligand complex and hepatocyte surface may be responsible for supplying more ligand to the cell for uptake. The surface-charged groups on albumin were modified to yield proteins having a range of isoelectric points (ALB, ALBs, ALBm, ALBe had values of 4.8-5.0, 4.5-4.7, 3.0-3.5, 8.4-8.6, respectively). [3H]-Palmitate uptake studies were performed with adult rat hepatocyte suspensions using similar unbound ligand fractions in the presence of the different binding proteins. Mass spectrometry, isoelectric focusing (pI), and heptane:water partitioning were used to determine protein molecular weight, pI, and protein-palmitate equilibrium binding constant, respectively. Hepatocyte [3H]-palmitate clearance in the presence of ALBs and ALBm were significantly lower (p < 0.05) than ALB, whereas [3H]-palmitate clearance in the presence of ALBe was significantly higher (p < 0.05) than ALB. The data were consistent with the notion that ionic interactions between extracellular protein-ligand complexes and the hepatocyte surface facilitate the uptake of long-chain fatty acids.

Albumins↗

PubMed2001

[1.4 kb 5' flanking region of class I patatin directs tuber-specific gus expression in potato (Solanum tuberosum L.)].

Binary vectors pPATIs (with partial signal sequence) and pPATI (without signal sequence) were constructed by fusing 1.4 kb 5' flanking regions of Class I patatin gene with GUS. Transient GUS expression was observed in in vitro tuber slices bombarded with pPATI. These constructs were then introduced into potato (cv. Desiree) via Agrobacterium tumefaciens transformation. Transgenic potato plants were confirmed by X-Gluc staining and PCR. Using in vitro tuberization system, GUS activities were assayed by fluorescence. It was shown that, in plants transformed with PATI-GUS, GUS specific activities were about 10-20 fold higher in tubers than in stems. Increased sucrose concentration could not induce PATI-GUS expression, but light enhanced PATI-GUS expression in cultured shoots.

Carboxylic Ester Hydrolases↗

PubMed2001

[30-year experiences on early detection and treatment of esophageal cancer in high risk areas].

The author had conducted the clinical study on early diagnosis and early therapy of esophageal cancer (EC) in the area of high prevalence for more than 30 years. In the endoscopic survey, 742 patients were diagnosed as early EC from 22,285 participants. The endoscopic appearance of the mucosa in early EC was classified into three types: (1) change of mucosa color: red and white area; (2) Increased thickness and change of the vascular pattern of the mucosa; (3) mucosa morphology: erosion, plaque, coarseness and nodular. Iodine staining under endoscopic examination had promoted the early detection of EC and precancerous lesions. Since 307 patients with EC at early stage (65 carcinoma in situ, 95 intramucosal carcinoma, and 147 submucosal carcinoma) had undergone esophagectomy. There were no lymphatic metastasis in carcinoma in situ, however there were 2 cases (2.11%) of intramucosal carcinoma and 27 cases (18.37%) of submucosal carcinoma with lymph node metastasis. 24-year follow-up, results showed that 5-year survival rate is 84.1% and a 10-year survival rate is 72.9% among the cases of esophagectomy. A 5-year survival rate is 100% in 22 cases followed up for full five years among 141 cases with endoscopic mucosectomy. According to this study, severe dysplasia and some moderate dysplasia were precancerous lesions. Should these precancerous cases be treated, the cancerization risk of target population would reduce dramatically and the incidence of EC would be decreased down to 2/3.

Adult↗

PubMed2000

Tumor-induced apoptosis of T cells: amplification by a mitochondrial cascade.

We have recently reported that apoptosis of T cells induced by squamous cell carcinoma of the head and neck (SCCHN) is partly Fas dependent. This tumor-induced T-cell death is mediated by the activities of caspase-8 and caspase-3 and is partially inhibited by antibodies to either Fas or Fas ligand. We report here that in contrast to apoptosis induced by agonistic anti-Fas antibody (Ab), the tumor-induced apoptotic cascade in Jurkat cells is significantly amplified by a mitochondrial loop. The involvement of mitochondria in tumor-induced apoptosis of T cells was demonstrated by changes in mitochondrial permeability transition as assessed by 3,3'-dihexiloxadicarbocyanine staining, by cleavage of cytosolic BID and its translocation to the mitochondria, by release of cytochrome c to the cytosol, and by the presence of active subunits of caspase-9 in Jurkat T cells cocultured with tumor cells. To further elucidate the significance of mitochondria in tumor-induced T-cell death, we investigated the effects of various inhibitors of the mitochondrial pathway. Specific antioxidants, as well as two inhibitors of mitochondria permeability transition, bongkrekic acid and cyclosporin A, significantly blocked the DNA degradation induced in Jurkat T cells by SCCHN cells. However, these inhibitors had no effect on cells triggered by anti-Fas Ab. Furthermore, a cell-permeable inhibitor of caspase-9, Ac-LEHD.CHO, which did not inhibit T-cell apoptosis induced by anti-Fas Ab, markedly inhibited apoptosis induced by etoposide or by coculture of Jurkat with SCCHN cells. These findings demonstrate that apoptotic cascades induced in Jurkat T lymphocytes by anti-Fas Ab or tumor cells are differentially susceptible to a panel of inhibitors of mitochondrial apoptotic events. It appears that besides the Fas-mediated pathway, additional mitochondria-dependent cascades are involved in apoptosis of tumor-associated lymphocytes. Inhibition of mitochondria-dependent cascades of caspase activation should be considered to enhance the success of immunotherapy or vaccination protocols in cancer.

Anti-Bacterial Agents↗

PubMed2000

Apoptosis-resistant mitochondria in T cells selected for resistance to Fas signaling.

Jurkat leukemic T cells are highly sensitive to the extrinsic pathways of apoptosis induced via the death receptor Fas or tumor necrosis factor-related apoptosis-inducing ligand as well as to the intrinsic/mitochondrial pathways of death induced by VP-16 or staurosporin. We report here that clonal Jurkat cell lines selected for resistance to Fas-induced apoptosis were cross-resistant to VP-16 or staurosporin. Each of the apoptotic pathways was blocked at an apical phase, where common regulators of apoptosis have not yet been defined. The Fas pathway was blocked at the level of caspase-8, whereas the intrinsic pathway was blocked at the mitochondria. No processing or activity of caspases was detected in resistant cells in response to either Fas-cross-linking or VP-16 treatment. Also, no apoptosis-associated alterations in the mitochondrial inner membrane, outer membrane, or matrix were detected in resistant Jurkat cells treated with VP-16. Thus, no changes in permeability transition, loss in inner membrane cardiolipin, generation of reactive oxygen species, or release of cytochrome c were observed in resistant cells treated with VP-16. Further, unlike purified mitochondria from wild type cells, those obtained from resistant cells did not release cytochrome c or apoptosis-inducing factor in response to recombinant Bax or truncated Bid. These results identify a defect in mitochondria ability to release intermembrane proteins in response to Bid or Bax as a mechanism of resistance to chemotherapeuetic drugs. Further, the selection of VP-16-resistant mitochondria via elimination of Fas-susceptible cells may suggest the existence of a shared regulatory component between the extrinsic and intrinsic pathways of apoptosis.

Antineoplastic Agents↗

PubMed2000

Inhibitor of apoptosis protein hILP undergoes caspase-mediated cleavage during T lymphocyte apoptosis.

Several endogenous or viral inhibitors of apoptosis, including Bcl-2, Bcl-xL, FLIP, p35, and CrmA, have been shown to be cleaved by caspases during apoptosis. In this study, we demonstrate that the endogenous inhibitor of apoptosis, hILP/XIAP, is also cleaved in apoptotic T lymphocytes, generating at least one prominent fragment of 29 kDa. This p29 cleaved fragment was detected in Jurkat cells induced to apoptose by anti-Fas antibody, staurosporin, or VP-16. The cleavage of hILP appears to be caspase mediated because the production of the p29 protein was inhibited by the pan-caspase peptide inhibitor, Z-VAD.FMK. In Jurkat cells engineered to overexpress CrmA, cleavage of hILP in response to anti-Fas antibody or staurosporin was inhibited, whereas overexpression of Bcl-2 abrogated the cleavage in response to VP-16. Cleavage of hILP was also observed in cell-free reactions using in vitro translated hILP and recombinant caspase-3 or -7. Moreover, we found that the p29 hILP fragment retained the ability to bind caspase-3 and -7, as shown previously for full-length or BIR-2 hILP. The p29 cleavage product was also detected during T-cell receptor-mediated apoptosis in peripheral blood lymphocytes from normal donors. Furthermore, tumor-associated T lymphocytes purified from ascites of patients with ovarian cancer expressed fragmented hILP, which was not detected in control T cells purified from peripheral blood of normal donors. Our results suggest that the cleavage of hILP represents an important event in apoptosis of T lymphocytes in both normal and pathological in vivo settings.

Amino Acid Chloromethyl Ketones↗

PubMed2000

Constitutive and inducible nitric oxide synthases after dynorphin-induced spinal cord injury.

It has recently been demonstrated that selective inhibition of both neuronal constitutive and inducible nitric oxide synthases (ncNOS and iNOS) is neuroprotective in a model of dynorphin (Dyn) A(1-17)-induced spinal cord injury. In the present study, various methods including the conversion of 3H-L-arginine to 3H-citrulline, immunohistochemistry and in situ hybridization are employed to determine the temporal profiles of the enzymatic activities, immunoreactivities, and mRNA expression for both ncNOS and iNOS after intrathecal injection of a neurotoxic dose (20 nmol) of Dyn A(1-17). The expression of ncNOS immunoreactivity and mRNA increased as early as 30 min after injection and persisted for 1-4 h. At 24-48 h, the number of ncNOS positive cells remained elevated while most neurons died. The cNOS enzymatic activity in the ventral spinal cord also significantly increased at 30 min 48 h, but no significant changes in the dorsal spinal cord were observed. However, iNOS mRNA expression increased later at 2 h, iNOS immunoreactivity and enzymatic activity increased later at 4 h and persisted for 24-48 h after injection of 20 nmol Dyn A(1-17). These results indicate that both ncNOS and iNOS are associated with Dyn-induced spinal cord injury, with ncNOS predominantly involved at an early stage and iNOS at a later stage.

Animals↗

PubMed2000

Antibiotic-heparin lock: in vitro antibiotic stability combined with heparin in a central venous catheter.

Long-term hemodialysis frequently requires vascular access through central venous catheters (CVCs). Infection related to these catheters is a significant complication. The use of an antibiotic-heparin lock could decrease the risks associated with infected permanent catheters. As an initial step in developing an antibiotic-heparin lock, we investigated the in vitro stability of antibiotic-heparin combinations in CVCs. Initially, cefazolin, vancomycin, ceftazidime, ciprofloxacin 10 mg/ml each, and gentamicin 5 mg/ml were incubated separately in glass test tubes in the dark at 37 degrees C for 72 hours. Samples were analyzed spectrophotometrically for stability at 24-hour intervals. The procedure was repeated with the addition of heparin (final concentration 5000 U/ml in glass test tubes), and the combination was also examined in CVCs. High-performance liquid chromatography analysis was conducted on the antibiotic-heparin combinations at 72 hours to confirm the spectrophotometric results. Ciprofloxacin produced an immediate precipitate with the addition of heparin and was not analyzed further. Absorbance values decreased for all antibiotics, with the greatest decreases at 72 hours for cefazolin (27.4%), vancomycin (29.7%), ceftazidime (40.2%), and gentamicin (8%) when combined with heparin. These decreases were postulated to be secondary to adsorption of the antibiotics to the luminal surface of the catheters because submitting the catheters to ultrasound with 1% sodium bicarbonate and analyzing the resulting solution for absorbance revealed that some of the drug was recovered. Although free antibiotic in CVC solution was reduced, the concentration should be sufficient (approximately 5 mg/ml) to decrease the frequency of infections associated with CVCs. We conclude that the concentrations of vancomycin, ceftazidime, cefazolin, or gentamicin used in our study should be sufficient for an antibiotic-heparin lock.

Anti-Bacterial Agents↗