Development and utility of the FDA 'GutProbe' DNA microarray for identification, genotyping and metagenomic analysis of commercially available probiotics.

AIM: Lactic acid bacteria are beneficial microbes added to many food products and dietary supplements for their purported health benefits. Proper identification of bacteria is important to assess safety as well as proper product labelling. A custom microarray (FDA GutProbe) was developed to verify accurate labelling in commercial dietary supplements. METHODS AND RESULTS: Strain-specific attribution was achieved with GutProbe array which contains genes from the most commonly found species in probiotic supplements and food ingredients. Applied utility of the array was assessed with direct from product DNA hybridization to determine (i) if identification of multiple strains in one sample can be conducted and (ii) if any lot-to-lot variations exist with eight probiotics found on the US market. CONCLUSIONS: GutProbe is a useful tool in identifying a mixture of microbials in probiotics and did reveal some product variations. In addition, the array is able to identify lot-to-lot differences in these products. These strain level attribution may be useful for routine monitoring of batch variation as part of a 'Good Manufacturing Practices' process. SIGNIFICANCE AND IMPACT OF THE STUDY: The FDA GutProbe is an efficient and reliable platform to identify the presence of microbial ingredients and determining microbe differences in dietary supplements. The GutProbe is a fast, rapid method for direct community profiling or food matrix sampling.

The inflammatory state of the rat placenta increases in late gestation and is further enhanced by glucocorticoids in the labyrinth zone.

INTRODUCTION: Inflammation plays central roles in key aspects of successful reproduction: ovulation, implantation and parturition. In this study we characterised the inflammatory profile of the rat placenta in late gestation with and without maternal glucocorticoid (dexamethasone) treatment. METHODS: Placentas (n = 6/group) were collected from untreated (Con) rats at days 16 and 22 (term = day 23) and from dexamethasone-treated (Dex) rats at day 22. mRNA and protein expression was determined for enzymes of prostaglandin synthesis and metabolism (Ptgs-1, Ptgs-2, 15-Pgdh), pro-inflammatory cytokines (Tnf-α, Il-1ß, Il-6), and the macrophage marker Emr-1 in the junctional (JZ) and labyrinth (LZ) zones of the placenta. RESULTS: Tnf-α, Il-1ß and Il-6 mRNAs all increased (2- to 4-fold) in both placental zones between days 16 and 22 (P < 0.01). Ptgs-2 mRNA (30-fold; P < 0.01) and PTGS-2 protein (2.4-fold; P < 0.05) similarly increased in LZ. In contrast, 15-Pdgh expression increased in JZ but decreased in LZ; these changes were accompanied by decreased levels of PGE2 in the JZ and a trend towards increased LZ levels. Dex treatment inhibited fetal and placental growth, but had minimal effects on expression of Ptgs-1, Ptgs-2 or 15-Pdgh. Nevertheless, Dex treatment increased LZ PGE2 levels (5-fold, P < 0.01) at the end of gestation. Dex treatment increased Tnf-α mRNA expression in LZ (40%; P < 0.05), but modestly suppressed cytokine protein expression in JZ. CONCLUSIONS: These data demonstrate that the inflammatory state of the LZ increases near term coincident with the known increase in local glucocorticoid levels. This suggests the classic anti-inflammatory actions of glucocorticoids do not occur in the placental LZ.

Kiss1 and Kiss1r mRNA expression in the rat placenta: changes with gestational age and regulation by glucocorticoids.

INTRODUCTION: Kisspeptin, the neuropeptide product of the KISS1 gene, is synthesized by neurons within the hypothalamus and is critical for fertility. Human placenta also expresses KISS1 and kisspeptin receptor (KISS1R) mRNA within the trophoblast compartment, where it is thought to act as a physiological invasion inhibitor. METHODS: We determined the expression of Kiss1 mRNA in rat placenta and examined the effect of gestational age and feto-placental growth restriction, achieved through excess maternal glucocorticoid exposure. RESULTS: Dexamethasone induced fetal growth restriction at both day 16 and day 22 of gestation, but placental growth restriction only at day 22. Real-time quantitative RT-PCR revealed an increase in Kiss1 and Kiss1r mRNA from day 16-22 in the labyrinth and junctional zones of the rat placenta. Immunolocalization confirmed kisspeptin expression in the placenta and was prominent in trophoblast tissue. Dexamethasone exposure elevated the expression of Kiss1 mRNA in the labyrinth and junctional zones of day 16 placentas. In contrast, Kiss1 mRNA in the labyrinth zone was reduced following dexamethasone-treatment at day 22. Kiss1r expression was increased in both placental zones at day 16 and 22 in response to dexamethasone-treatment. CONCLUSIONS: We confirm the presence of Kiss1 and Kiss1r mRNA in the rat placenta with expression increasing over the final third of pregnancy, suggestive of a role in restricting placental growth. Furthermore, the effects of dexamethasone on placental Kiss1/Kiss1r suggest glucocorticoid-induced placental growth retardation could be mediated, in part, via early stimulation of Kiss1 and the subsequent inhibition of trophoblast proliferation and invasion.

Impact of five commercial baits containing chitin synthesis inhibitors on the protist community in Reticulitermes flavipes (Isoptera: Rhinotermitidae).

Eastern subterranean termite, Reticulitermes flavipes (Kollar), workers were continuously exposed to one of five chitin synthesis inhibiting (CSI) active ingredients and the protist community from the hindgut quantified biweekly for 21 d. The CSIs tested included commercially available formulations of diflubenzuron, hexaflumuron, lufenuron, noviflumuron, and novaluron. Results showed termites exposed to CSIs had a significant decrease (>or=30%) in the estimated total protist population after 3 d, regardless of treatment. Protist species impacted were Dinenympha fimbriata, D. gracilis, Microjoenia fallax, Pyrsonympha vertens, and Trichonympha agilis and could be indicative of weakened digestive homeostasis, but further studies are needed. We also provide evidence that lufenuron is highly toxic and discuss some of the implications this might have on termite management practices.

Effect of dermatan sulfate on the indentation and tensile properties of articular cartilage.

OBJECTIVE: This paper examines the hypothesis that the dermatan sulfate (DS) chain on decorin is a load carrying element in cartilage and that its damage or removal will alter the material properties. METHODS: To test this hypothesis, indentation and tensile testing of cartilage from bovine patella were performed before and after digestion with chondroitinase B (cB). Removal of significant amounts of DS by cB digestion was verified by Western blot analysis of proteoglycans extracted from whole and sectioned specimens. Specimens (control and treated) were subjected to a series of step-hold displacements. Elastic modulus during the step rise (rapid modulus) and at equilibrium (equilibrium modulus), and the relaxation function during each step was measured for test (cB and buffer) and control (buffer alone) conditions. RESULTS: cB had no effect on any of the viscoelastic mechanical properties measured, either in indentation or tension. CONCLUSION: Removing or damaging approximately 50% of the DS had no effect on the mechanical properties, strongly suggesting that DS either carries very low load or no load.

Evaluation of fracture toughness of cartilage by micropenetration.

Failure properties of cartilage are important in injury repair and disease, but few methods exist for measuring these properties, especially in small animals. To meet this need, a new indentation/penetration method for measuring fracture toughness of cartilage is proposed. During indentation, a conical tip is displaced into the surface of the cartilage, causing first a non-penetrating indentation, and then a penetration into the tissue. The method assumes that tissue penetration occurs during periods of "rapid work", which are identified from a curve of work rate vs. time. Total penetration depth is determined by summing the displacement during these periods. Fracture work is the work that occurs during "rapid work", or penetration, and fracture toughness defined as the fracture work divided by one-half the penetrated surface area of the indenting tip. The method was validated by indentation testing of bovine cartilage. Penetrating indentations with a conical tip were performed in bovine patellar cartilage and depth of penetration and fracture toughness predicted. For comparison with the indentation data, depth of penetration was measured in histological sections. These measurements agreed well with the predicted depth. Predicted fracture toughness also agreed with values measured via a macroscopic test. This newly described method has promise as a general method for measuring fracture toughness in cartilage, particularly in small animals, since penetrating tips with small tip radius can be manufactured and penetration may be accomplished in cartilage of minimal thickness.

Phosphatidylinositol-3 kinase inhibitors reproduce the selective antiproliferative effects of imatinib on chronic myeloid leukaemia progenitor cells.

We investigated the role of the phosphatidylinositol-3 kinase (PI-3K) pathway in regulating the proliferation of primary chronic myeloid leukaemia (CML) progenitor cells by using imatinib to inhibit the activity of p210(Bcr-Abl). The effect of imatinib on the expression of PI-3K pathway proteins was investigated by kinase assays and Western blotting; PI-3K was inhibited by wortmannin or LY294002, Jak2 by AG490 and farnesylation by FTI II; progenitor cell proliferation (self-renewal) was measured by growing myeloid colonies in vitro, then replating them to observe secondary colony formation. Suppression of p210(Bcr-Abl) with imatinib indirectly suppressed the activity of PI-3K and its downstream targets (Erk, Akt and p70S6 kinase), thereby implicating the PI-3K pathway in p210(Bcr-Abl)-mediated signalling in primary CML progenitor cells. The PI-3K inhibitors, wortmannin and LY294002 reproduced the differential effects of imatinib on normal and CML progenitor cell proliferation in vitro by increasing normal cell (P = 0.001) and reducing CML cell proliferation (P = 0.0003). This differential effect was attributable to dysregulated signalling by granulocyte colony-stimulating factor in CML. The responses of individual patient's cells to wortmannin correlated with their responses to imatinib (P = 0.004) but not their responses to AG490 (Jak2 kinase inhibitor) or FTI II (farnesyltransferase inhibitor). Individual responses to wortmannin also correlated with responses to interferon alpha (IFNalpha) (P = 0.016). Imatinib-resistant K562 cells were sensitive to LY294002. Inhibition of the PI-3K pathway may be common to imatinib and IFNalpha and reflect dysregulated cytokine signalling. As imatinib-resistant cells remained sensitive to wortmannin and LY294002, targeting the PI-3K pathway may provide an alternative therapy for imatinib-resistant patients.

Effect of specimen thickness on fracture toughness of bovine patellar cartilage.

Fracture toughness and crack tip opening angle were measured for bovine patellar cartilage using modified single-edged notch specimens of two thicknesses. There was no difference in fracture toughness between thin (0.7 mm) versus relatively thick (2.7 mm) specimens, but the crack tip opening angle at initiation of crack propagation was larger for the thin specimens (106 deg) than for the thick specimens (70 deg). Fracture toughness of the bovine patellar cartilage (1.03 kJ/m2) was not statistically different than that reported previously for canine patellar cartilage (1.07 kJ/m2) employing the same methods. Large variation in measurements for both bovine and canine cartilage are in part attributable to variation between individual animals, and are consistent with variation in other mechanical property measurements for articular cartilage. The observed reduction in crack tip opening angle with increased specimen thickness is consistent with behavior of some engineering materials, and demonstrates that specimen thickness influences fracture behavior for bovine patellar cartilage.

Collagen architecture and failure processes in bovine patellar cartilage.

Cartilage fails by fibrillation and wearing away. This study was designed to identify the microscopic failure processes in the collagen network of bovine cartilage using scanning electron microscopy. Cartilage samples from fibrillated cartilage from the bovine patella were removed from the bone, fixed, digested to remove proteoglycans, freeze-fractured, and processed for SEM. The architecture of the collagen network in the normal cartilage was first defined, and then the failure processes were identified by examining sites of fibrillation and at crack tips. The bovine patellar cartilage was organised with a superficial layer composed of 3-5 lamina, attached to a sub-superficial tissue by angled bridging fibrils. Collagen in the sub-superficial tissue was organised in lamina oriented in the radial direction up to the transition zone. Failure of the system occurred by cracks forming in superficial layer and lamina, creating flaps of lamina that rolled up into the larger 'fronds'. Larger cracks not following the laminar planes occurred in the transition, mid, and deep zones. Failure at the crack tips in the sub-superficial tissue appeared to be by peeling of collagen fibrils, as opposed to breaking of collagen fibrils, suggesting a 'glue' bonding the collagen fibrils in a parallel fashion. Cracks propagated by breaking these bonds. This bond could be a site of disease action, since weakening of the bond would accelerate crack propagation.

The benefit of a special elective gynecologic oncology program for obstetrics and gynecology residents.

OBJECTIVE: To assess the benefit of a special elective gynecologic oncology program for Obstetrics and Gynecology (Ob/Gyn) residents. METHODS: We reviewed our housestaff records from July 1992 to June 1998 and the National Residency Matching Program (NRMP) subspeciality match results for gynecologic oncology from its inception in 1994 to 1999. RESULTS: From July 1992 to June 1998, a total of 146 residents participated in our elective program. Of the 104 candidates who went through our program and subsequently participated in the NRMP, 55 (53%) obtained match positions. After completion of the elective, 42 of the 146 residents (29%) did not participate in the NRMP for gynecologic oncology and therefore were not eligible to obtain match appointments. During the study period, there were 255 other residents in the United States who applied for gynecologic oncology fellowship positions through the NRMP and did not participate in our program. Of these 255 candidates, 137 (54%) matched. CONCLUSION: The percentage of residents who went through our program, participated in the NRMP, and obtained fellowships did not differ significantly from the percentage of residents who matched without participating in the program. However, almost one-third of the residents who went through our program did not participate in the NRMP. The reasons for their lack of participation were not formally evaluated, but are likely related to a personal decision to pursue another carrer pathway, a decision facilitiated by their experience in our program. Therefore, it appears that the main benefits of the program are to help potential candidates decide whether or not to pursue a career in gyencologic oncology and to aid fellowship programs in identifying exceptional candidates for subspecialty training.

Progenitor cells from patients with advanced phase chronic myeloid leukaemia respond to STI571 in vitro and in vivo.

STI571 targets p210(BCR-ABL) in chronic myeloid leukaemia (CML). In vitro, STI571 reduces self-replication (replating ability) by chronic-phase CML CFU-GM. Here, we studied CFU-GM in advanced-phase (accelerated and blast crisis) CML. The numbers and self-replication of CFU-GM in advanced phase were greater than in the chronic phase. Self-replication by CFU-GM from advanced phase patients was reduced by STI571 or IFN alfa to the same extent as in the chronic phase. The reduced replating ability induced by STI571 correlated with that induced by IFN alpha (r=0.73). STI571 treatment in vivo also reduced replating ability and the numbers of CFU-GM/ml of blood.

Depression and blood types.

A sample of 108 normal volunteers (grouped as blood Types O, A, and B/AB) were administered the Beck Depression Inventory. The results suggest that the association between depression and blood Type O that has been found for hospitalized patients can also be observed in normal patients.

Olfactory bulb uptake and determination of biotransfer factors in the California ground squirrel (Spermophilus beecheyi) exposed to manganese and cadmium in environmental habitats.

Manganese (Mn) and cadmium (Cd) profiles in olfactory bulbs of California ground squirrels (Spermophilus beecheyi) trapped at Lawrence Livermore National Laboratory's Site 300 facility in California were determined with proton induced X-ray emission (PIXE). As a reference, Mn profiles in olfactory bulbs from laboratory rats exposed via nose-only inhalation to 0.53 mg/m3 Mn in the form of MnCl2 were also determined with PIXE. Atomic absorption spectrophotometry was used to measure soil Mn and Cd contents from the trapping sites and Mn and Cd contents in ground squirrel liver and leg muscle tissues. The data from laboratory rats revealed that Mn uptake into the olfactory bulb occurs via inhalation exposure. Data from ground squirrels and knowledge of the collection sites indicate that although several routes of exposure may occur, fossorial rodent olfactory uptake affords a significant exposure route to Mn and Cd in soils. Measured biotransfer factors (ratio of leg muscle tissue metal content to soil metal content) for Cd in ground squirrels were 10(3)-fold greater than exposure modeling estimates based on oral Cd uptake data from livestock. The measurements for ground squirrel tissues show that when conducting ecological risk assessments for natural habitats considerable care should be taken in selecting transfer factors. Specifically, transfer factors derived from data pertaining to comparable exposure pathways and ecological setting should be used wherever possible.

Neoadjuvant therapy for adenocarcinoma of the rectum: tumor response and acute toxicity.

PURPOSE: This study was designed to evaluate the down-staging effect and acute toxicity of preoperative radiation and chemoradiation for primary adenocarcinoma of the rectum. METHODS: The results of pretreatment staging with transrectal ultrasound and computed tomography were compared with final histologic stage in 260 consecutive patients who underwent neoadjuvant therapy and proctectomy for primary adenocarcinoma of the rectum. Patients underwent short-course radiation (2,000 cGy in five fractions), long-course radiation (4,500 cGy in 25 fractions), or chemoradiation (4,500 cGy in 25 fractions with concurrent chemotherapy). RESULTS: Down-staging of one or more T stages occurred in 116 of 260 (45 percent) patients overall (short-course radiation 34/82 (42 percent), long-course radiation 55/122 (45 percent), chemoradiation 27/56 (48 percent), P = not significant). Down-staging of one or more N stages occurred in 85 of 178 (48 percent) patients overall (short-course radiation 12/45 (27 percent), long-course radiation 49/86 (57 percent), chemoradiation 24/47 (51 percent), P = 0.003). Complete pathologic response was observed in 16 of 260 (6 percent) patients overall (short-course radiation 4/82 (5 percent), long-course radiation 5/122 (4 percent), chemoradiation 7/56 (13 percent), P = 0.08). Resection with negative margins (distal, proximal, and radial) was achieved in 211 of 227 patients (93 percent) in whom complete radial margin data were available. Permanent stomas were created in 35 percent of patients; temporary stomas were created in 15 percent. Thirty-three Grade 3 or 4 toxicities occurred in 22 of 260 (8 percent) patients overall during neoadjuvant therapy. Toxicity was more frequent in patients receiving chemoradiation (14/56; 25 percent) and long-course radiation (8/122; 7 percent) than in those receiving short-course radiation (0/82; 0 percent), P < 0.0001. Perioperative complications occurred in 93 patients overall (36 percent). The postoperative mortality rate was 0.4 percent (1/260). There was no significant difference in the complication rate between patients treated with short-course radiation (26/82; 32 percent), long-course radiation (46/122; 36 percent), and chemoradiation (21/56; 38 percent). CONCLUSION: Neoadjuvant therapy for adenocarcinoma of the rectum is well tolerated and can produce substantial down-staging and a high curative resection rate. Chemoradiation can achieve high complete pathologic response rates, although toxicity during neoadjuvant therapy is greater than for radiation alone. Short-course radiation can achieve down-staging of both T stage and N stage.

The influence of INK4 proteins on growth and self-renewal kinetics of hematopoietic progenitor cells.

This study investigated the influence of expression of proteins of the INK4 family, particularly p16, on the growth and self-renewal kinetics of hematopoietic cells. First, retrovirus-mediated gene transfer (RMGT) was used to restore p16(INK4a) expression in the p16(INK4a)-deficient lymphoid and myeloid cell lines BV173 and K562, and it was confirmed that this inhibited their growth. Second, to sequester p16(INK4a) and related INK4 proteins, cyclin-dependent kinase 4 (CDK4) was retrovirally transduced into normal human CD34(+) bone marrow cells and then cultured in myeloid colony-forming cell (CFC) assays. The growth of CDK4-transduced colonies was more rapid; the cell-doubling time was reduced; and, upon replating, the colonies produced greater yields of secondary colonies than mock-untransduced controls. Third, colony formation was compared by marrow cells from p16(INK4a-/-) mice and wild-type mice. The results from p16(INK4a-/-) marrow were similar to those from CDK4-transduced human CFCs, in terms of growth rate and replating ability, and were partially reversed by RMGT of p16(INK4a). Lines of immature granulocytic cells were raised from 15 individual colonies grown from the marrow of p16(INK4a-/-) mice. These had a high colony-forming ability (15%) and replating efficiency (96.7%). The p16(INK4a-/-) cell lines readily became growth factor-independent upon cytokine deprivation. Taken together, these results demonstrate that loss of INK4 proteins, in particular p16(INK4a), increases the growth rate of myeloid colonies in vitro and, more importantly, confers an increased ability for clonal expansion on hematopoietic progenitor cells.

BCR-ABL and interleukin 3 promote haematopoietic cell proliferation and survival through modulation of cyclin D2 and p27Kip1 expression.

Although it is evident that BCR-ABL can rescue cytokine-deprived hematopoietic progenitor cells from cell cycle arrest and apoptosis, the exact mechanism of action of BCR/ABL and interleukin (IL)-3 to promote proliferation and survival has not been established. Using the pro-B cell line BaF3 and a BaF3 cell line stably overexpressing BCR-ABL (BaF3-p210), we investigated the proliferative signals derived from BCR-ABL and IL-3. The results indicate that both IL-3 and BCR-ABL target the expression of cyclin Ds and down-regulation of p27(Kip1) to mediate pRB-related pocket protein phosphorylation, E2F activation, and thus S phase progression. These findings were further confirmed in a BaF3 cell line (TonB.210) where the BCR-ABL expression is inducible by doxycyclin and by using the drug STI571 to inactivate BCR-ABL activity in BaF3-p210. To establish the functional significance of cyclin D2 and p27(Kip1) expression in response to IL-3 and BCR-ABL expression, we studied the effects of ectopic expression of cyclin D2 and p27(Kip1) on cell proliferation and survival. Our results demonstrate that both cyclin D2 and p27(Kip1) have a role in BaF3 cell proliferation and survival, as ectopic expression of cyclin D2 is sufficient to abolish the cell cycle arrest and apoptosis induced by IL-3 withdrawal or by BCR-ABL inactivation, while overexpression of p27(Kip1) can cause cell cycle arrest and apoptosis in the BaF3 cells. Furthermore, our data also suggest that cyclin D2 functions upstream of p27(Kip1), cyclin E, and cyclin D3, and therefore, plays an essential part in integrating the signals from IL-3 and BCR-ABL with the pRB/E2F pathway.

Peripheral blood progenitor cell mobilisation alters myeloid, but not erythroid, progenitor cell self-renewal kinetics.

Transplantation of progenitor cells which have been mobilised into the bloodstream (PBPC) following the administration of G-CSF results in more rapid neutrophil recovery than transplantation of bone marrow (BM). The reasons for the accelerated neutrophil engraftment are not clear, but would be explained by increased self-replication of myeloid progenitor cells (CFU-GM). We have used a CFU-GM replating assay to investigate myeloid progenitor self-replication, and quantification of subcolony formation during erythroid burst formation to quantify erythroid progenitor self-renewal. Secondary colony formation by CFU-GM, grown from PBPC and then replated was increased compared with secondary colony formation by BM CFU-GM (P = 0.0001); erythroid subcolony formation was not altered. There was no difference between the replating abilities of PBPC CFU-GM derived from allogeneic donors (normal individuals) and autologous donors (patients with malignant disease) although differences were found between subgroups of autologous donors. The increased replication of PBPC could not be accounted for by a reduction in progenitor cell apoptosis; PBPC CFU-GM contained slightly fewer apoptotic CD34+ cells than BM CFU-GM. The increased replication by PBPC CFU-GM was reversible because it declined when CFU-GM colonies were passaged through three sequential CFU-GM replating cycles. This decline in self-replication was more rapid than the decline seen in replated BM CFU-GM. The self-replication of PBPC CFU-GM, and subcolony formation by BFU-E could be further enhanced by exposure to cytokines in vitro. We conclude that mobilisation alters the replication kinetics of myeloid, but not of erythroid, progenitor cells, that mobilisation-induced events are of limited duration and that in vitro exposure to cytokines may modify PBPC progenitor cell kinetics.

The transmembrane domain of syntaxin 1A is critical for cytoplasmic domain protein-protein interactions.

Assembly of the plasma membrane proteins syntaxin 1A and SNAP-25 with the vesicle protein synaptobrevin is a critical step in neuronal exocytosis. Syntaxin is anchored to the inner face of presynaptic plasma membrane via a single C-terminal membrane-spanning domain. Here we report that this transmembrane domain plays a critical role in a wide range of syntaxin protein-protein interactions. Truncations or deletions of the membrane-spanning domain reduce synaptotagmin, alpha/beta-SNAP, and synaptobrevin binding. In contrast, deletion of the transmembrane domain potentiates SNAP-25 and rbSec1A/nsec-1/munc18 binding. Normal partner protein binding activity of the isolated cytoplasmic domain could be "rescued" by fusion to the transmembrane segments of synaptobrevin and to a lesser extent, synaptotagmin. However, efficient rescue was not achieved by replacing deleted transmembrane segments with corresponding lengths of other hydrophobic amino acids. Mutations reported to diminish the dimerization of the transmembrane domain of syntaxin did not impair the interaction of full-length syntaxin with other proteins. Finally, we observed that membrane insertion and wild-type interactions with interacting proteins are not correlated. We conclude that the transmembrane domain, via a length-dependent and sequence-specific mechanism, affects the ability of the cytoplasmic domain to engage other proteins.

Initial experience of a nephrologist-operated vascular access center.

A nephrology practice in Alabama did not feel in control of vascular access management. Scheduling delays, as well as variable techniques and outcomes, leading to high morbidity and mortality, caused frustration with the existing care system for vascular access. Our objective was to develop an integrative system of vascular access care, involving nephrologists along with the other caregivers, and to demonstrate an improvement in outcomes. Nephrology Vascular Labs (NVL), a recent RMS-Lifeline acquisition, opened a vascular access center (VAC) as an extension of the nephrology practice. Both pre-ESRD and ESRD patients are evaluated and treated in the VAC. Treatment is rendered in a timely fashion, to the benefit of the patients. Nephrologists serve as the interventionists. More than 90% of vascular access problems detected at dialysis are treated at the VAC. More than 2000 procedures have been performed over 2 years. Procedures carried out include thrombolysis with angioplasty, fluoroscopy alone or with angioplasty, placement of cuffed and noncuffed catheters, removal of cuffed catheters, and minor surgeries. Success rates have been high. Minor and major complications have been relatively low. Referrals to both surgeons and radiologists are shown to emphasize the role of the VAC as part of an integrative system of vascular access care. Results of a patient satisfaction survey were excellent. The VAC has fulfilled the vision of creating a seamless integration of care for vascular access. Hospitalization rate has been reduced and it is suspected that the global cost of access care is markedly lower than prior to the VAC. Multiple nephrologists can rotate as the VAC's interventionist and jointly obtain good outcomes and have little variability among them. Several reasons for using a nephrologists as the interventionist are discussed.

A simple cryopreservation method for the maintenance of cell viability and mechanical integrity of a cultured cartilage analog.

A method for cryopreserving a 100-microm-thick sheet of tissue produced by cultured rabbit chondrocytes has been developed. The method maintains cell viability and avoids tissue fracture and degradation of mechanical properties. A slow-freeze, fast-thaw procedure with 2 M Me(2)SO as the cryoprotectant resulted in no tissue fracture and approximately 90% viable cells after storage in culture flasks at -80 degrees C. The cells in the retrieved tissue remained responsive to IL-1beta, and tensile and fracture toughness properties of the tissue were not degraded by cryopreservation.