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Biomedical subjects

S L Woo

Publications and source records attributed to S L Woo.

At least 73 records · Page 4Linked to original sources
PubMed1999

Expression of two major chitinase genes of Trichoderma atroviride (T. harzianum P1) is triggered by different regulatory signals.

Regulation of the expression of the two major chitinase genes, ech42 (encoding the CHIT42 endochitinase) and nag1 (encoding the CHIT73 N-acetyl-beta-D-glucosaminidase), of the chitinolytic system of the mycoparasitic biocontrol fungus Trichoderma atroviride (= Trichoderma harzianum P1) was investigated by using a reporter system based on the Aspergillus niger glucose oxidase. Strains harboring fusions of the ech42 or nag1 5' upstream noncoding sequences with the A. niger goxA gene displayed a glucose oxidase activity pattern that was consistent under various conditions with expression of the native ech42 and nag1 genes, as assayed by Northern analysis. The expression product of goxA in the mutants was completely secreted into the medium, detectable on Western blots, and quantifiable by enzyme-linked immunosorbent assay. nag1 gene expression was triggered during growth on fungal (Botrytis cinerea) cell walls and on the chitin degradation product N-acetylglucosamine. N-Acetylglucosamine, di-N-acetylchitobiose, or tri-N-acetylchitotriose also induced nag1 gene expression when added to mycelia pregrown on different carbon sources. ech42 expression was also observed during growth on fungal cell walls but, in contrast, was not triggered by addition of chitooligomers to pregrown mycelia. Significant ech42 expression was observed after prolonged carbon starvation, independent of the use of glucose or glycerol as a carbon source, suggesting that relief of carbon catabolite repression was not involved in induction during starvation. In addition, ech42 gene transcription was triggered by physiological stress, such as low temperature, high osmotic pressure, or the addition of ethanol. Four copies of a putative stress response element (CCCCT) were found in the ech42 promoter.

Aspergillus niger↗
PubMed1999

Molecular correlations in phenylketonuria: mutation patterns and corresponding biochemical and clinical phenotypes in a heterogeneous California population.

We studied 133 California phenylketonuria (PKU) patients and one obligate heterozygote to delineate the molecular basis of PKU in a population with greater ethnic diversity than in previous studies, and to determine whether a correlation exists between genotype and clinical phenotype, with the latter defined by both the diagnostic pretreatment blood phenylalanine (PHE) level and cognitive (IQ) test scores. To determine PAH genotypes, we used PCR-mediated amplification, denaturing gradient gel electrophoresis, and direct sequencing on dried whole blood samples. Where possible, mutation severity was defined according to predicted in vitro PAH enzyme activity estimated by using Cos cell expression analysis for a given mutation. We then asked whether mutation severity, as defined by such expression analysis, correlated with pretreatment PHE levels or with IQ test results. A mutation was identified in 236 (88%) of 267 mutant alleles. Seventeen new mutant alleles were found; A47E, T81P, I102T, E182G, T328D, Y343P, K371R, Y387H, A389E, E422K, IVS9nt5, IVS11nt20, delS70, del364-368/del198-220, delF299, delT323, and -1C/T. In striking contrast to a number of studies in other populations, in this study, based on predicted PAH activity, we observed no correlation between mutation severity and pretreatment PHE levels. There was also no correlation between genotype and IQ. We conclude that in samples collected from an ethnically heterogeneous population, there is no correlation of mutation severity with either pretreatment PHE levels or IQ measurement in treated patients. We caution that genetic counseling in PKU should incorporate the notion that prognosis may not be predicted with precision based on mutation analysis in a given patient.

California↗
PubMed1998

Genes from mycoparasitic fungi as a source for improving plant resistance to fungal pathogens.

Disease resistance in transgenic plants has been improved, for the first time, by the insertion of a gene from a biocontrol fungus. The gene encoding a strongly antifungal endochitinase from the mycoparasitic fungus Trichoderma harzianum was transferred to tobacco and potato. High expression levels of the fungal gene were obtained in different plant tissues, which had no visible effect on plant growth and development. Substantial differences in endochitinase activity were detected among transformants. Selected transgenic lines were highly tolerant or completely resistant to the foliar pathogens Alternaria alternata, A. solani, Botrytis cinerea, and the soilborne pathogen Rhizoctonia solani. The high level and the broad spectrum of resistance obtained with a single chitinase gene from Trichoderma overcome the limited efficacy of transgenic expression in plants of chitinase genes from plants and bacteria. These results demonstrate a rich source of genes from biocontrol fungi that can be used to control diseases in plants.

Base Sequence↗
PubMed1998

Retrovirus-mediated in vivo gene transfer in the replicating liver using recombinant hepatocyte growth factor without liver injury or partial hepatectomy.

Retrovirus-mediated gene delivery into hepatocytes in vivo provides long-term gene expression, which is of great importance for treating most genetic and metabolic disorders. However, clinical application has not been realized because of the requirement for prior 70% partial hepatectomy or chemical (toxic) liver injury to initiate hepatocyte replication at the time of retroviral gene transduction. In this paper, we describe a novel gene delivery system that uses recombinant hepatocyte growth factor (rHGF) prior to retrovirus-mediated in vivo gene transfer in the liver without partial hepatectomy or liver injury. A single retroviral infusion through the portal vein following five systemic injections (via the tail vein) of 100 microg/kg rHGF resulted in a 10.4% 5-bromo-2'-deoxyuridine (BrdU) labeling index (BLI) and 0.14% retroviral gene transduction efficiency (RGTE) in hepatocytes, which were 6.3- and 12.9-fold higher than those of controls, respectively. Modest additional increases in BLI and RGTE (13.4% and 0.22%, respectively) were seen after five systemic injections of 500 microg/kg rHGF. The correlation between BLI and RGTE was statistically confirmed regardless of treatment. When rats received multiple retroviral infusions through a cannulated portal vein following five portal injections of 100 microg/kg rHGF, RGTE was dramatically increased (1.3%) and in some areas of the liver exceeded more than 10%. There was no evidence of liver injury in any animal. This approach has great potential for clinical application in terms of avoiding invasive procedures or liver injury.

Animals↗
PubMed1998

Quantitative anthropometry of the subatlantal cervical longitudinal ligaments.

STUDY DESIGN: The quantitative anthropometry of the cervical longitudinal ligaments was determined in 20 human cadaveric subatlantal cervical spines at the limits of flexion and extension. OBJECTIVES: To provide measurements of cervical anterior and posterior longitudinal ligament lengths, widths, and cross-sectional areas at segmental levels. SUMMARY OF BACKGROUND DATA: Although mathematical models of the cervical spine require specific data to predict kinematics, the anthropometry of the cervical spine has not been examined in detail. The dimensional changes of ligaments in physiologic motion are not well characterized. METHODS: Segmental lengths and widths of the cervical longitudinal ligaments were measured in sagittal plane flexion and extension, using a three-dimensional electromagnetic digitizer. The cross-sectional areas of the ligaments at resting length were measured with a laser micrometer system. Comparisons between anterior and posterior location and among segmental levels were made. Several ligaments were examined histologically to determine the insertion sites and, thus, to define the segmental length. RESULTS: The anterior longitudinal ligaments were shorter in flexion than in extension. In extension, they were longer than the posterior longitudinal ligaments in flexion. The resting isolated ligaments were longer than the longest in situ lengths at several vertebral levels. The anterior longitudinal ligaments were wider at the disc than at the body. The cross-sectional area at C2-C3 was smaller than at subaxial levels. The longitudinal ligaments were observed to insert along the entire underlying vertebral body. CONCLUSIONS: The quantitative anthropometry of the cervical longitudinal ligaments is important in the development of accurate mathematical models of the cervical spine. The in situ ligaments may not be under tension in the physiologic range of motion.

Aged↗
PubMed1998

Effect of combined axial compressive and anterior tibial loads on in situ forces in the anterior cruciate ligament: a porcine study.

This study investigated the impact of a combination of axial compressive and anterior-posterior tibial loads on the in situ forces in the anterior cruciate ligament. An axial compressive load is believed to contribute to increased stability of the knee joint; however, its effect on in situ forces in the anterior cruciate ligament has not been clearly defined, to our knowledge. It was hypothesized that the application of an axial compressive load, when combined with an anterior tibial load, would result in larger in situ forces in the anterior cruciate ligament than those caused by an isolated anterior tibial load. With use of a porcine knee model, the results confirmed this hypothesis; the addition of a 200 N axial compressive load to a 100 N anterior tibial load increased knee stability by reducing anterior-posterior tibial translation and internal-external tibial rotation and also caused a significant increase in in situ forces in the anterior cruciate ligament (p < 0.05). Specifically, there was a 34% increase in the in situ force at 30 degrees of flexion, a 68% increase at 60 degrees of flexion, and an 84% increase at 90 degrees of flexion compared with those for an isolated anterior tibial load of 100 N. Additionally, there was a statistically significant increase of the in situ forces in the anterior cruciate ligament at 60 and 90 degrees as compared with those at 30 degrees. These results suggest that axial compressive loads on the knee may play a role in injury of the anterior cruciate ligament when the knee is flexed.

Animals↗
PubMed1998

In situ forces in the posterolateral structures of the knee under posterior tibial loading in the intact and posterior cruciate ligament-deficient knee.

The posterolateral structures of the knee consist of a complex anatomical architecture that includes several components with both static and dynamic functions. Injuries of the posterolateral structures occur frequently in conjunction with ruptures of the posterior cruciate ligament. To investigate the role of the posterolateral structures in maintaining posterior knee stability, we measured the in situ forces in the posterolateral structures and the distribution of force within the structures' major components, i.e., the popliteus complex and the lateral collateral ligament, in response to a posterior tibial load. Eight cadaveric knees were tested. With use of a robotic/universal force-moment sensor testing system, a posterior tibial load of 110 N was applied to the knee, and the resulting five-degree-of-freedom kinematics were measured at flexion angles of 0, 30, 60, 75, and 90 degrees. The knees were tested first in the intact state and then after the posterior cruciate ligament had been resected. These tests were also performed with an additional load of 44 N applied at the aponeurosis to simulate contraction of the popliteus muscle. In the intact knee, the in situ forces in the posterolateral structures were found to decrease with increasing knee flexion. After the posterior cruciate ligament was sectioned, these forces increased significantly at all angles of flexion. With no load applied to the popliteus muscle, the in situ forces in the popliteus complex were similar to those in the lateral collateral ligament. However, with a load of 44 N applied to the popliteus muscle, in situ forces in the popliteus complex were three to five times higher than those in the lateral collateral ligament. These results reveal that in response to posterior tibial loads, the posterolateral structures play an important role at full extension in intact knees and at all angles of flexion in posterior cruciate ligament-deficient knees. The popliteus muscle appears to be a major stabilizer under this loading condition; thus, the inability to restore its function may be a cause of unsatisfactory results in reconstructive procedures of the posterolateral structures of the knee.

Aged↗
PubMed1998

Engineering the healing of the rabbit medial collateral ligament.

A biological approach to improve healing of the medial collateral ligament (MCL) was investigated by exploring the use of therapeutic growth factors based on in vitro and in vivo experiments. The in vitro cell culture studies involved screening a variety of growth factors to select those that exhibit the most positive effects on cell proliferation and extracellular matrix synthesis. The selected growth factors were applied in vivo to a rabbit model where the MCL was ruptured. Biomechanical and histological evaluations are performed to determine whether the selected growth factors can enhance the properties of the healed MCL, whether these improvements are dose dependent, and whether combinations of growth factors can enhance MCL healing to a greater extent than individual growth factors. In vitro studies showed that epidermal growth factor (EGF) and platelet derived growth factor-BB (PDGF-BB) have the greatest effect on ligament fibroblast proliferation, whereas transforming growth factor-beta 1 (TGF-beta 1) superiorly promotes extracellular matrix synthesis. These growth factors were then applied in vivo at different dosages, in isolation and in combination, and the ligaments were evaluated six weeks post-operatively. Tensile testing of the femur-MCL-tibia complexes (FMTCs) revealed that the specimens treated with a high dose of PDGF-BB have ultimate load, ultimate elongation and energy absorbed to failure values that are significantly greater than those from the other groups. The high dose of PDGF-BB was more effective than the low dose, indicating a dose dependency. The addition of TGF-beta 1 to PDGF-BB did not lead to any further increases in the structural properties of the FMTC. These encouraging results suggest that PDGF-BB may be a potential growth factor to enhance the quality of the healing ligament.

Animals↗
PubMed1998

The effects of multiple-strand suture methods on the strength and excursion of repaired intrasynovial flexor tendons: a biomechanical study in dogs.

This study was designed to determine the effects of in vivo multistrand, multigrasp suture techniques on the strength and gliding of repaired intrasynovial tendons when controlled passive motion rehabilitation was used. Twenty-four adult mongrel dogs were divided into 4 groups and their medial and lateral forepaw flexor tendons were transsected and sutured by either the Savage, the Tajima, the Kessler, or the recently developed 8-strand suture method. The tendon excursion, joint rotation, and tensile properties of the repaired tendons were evaluated biomechanically at 3 and 6 weeks after surgery. It was found that neither time nor suture method significantly effected proximal and distal interphalangeal joint rotation or tendon excursion when the 4 techniques were compared to each other. Normalized load value (experimental/control) was significantly affected by both the suture method and the amount of time after surgery, however. The Savage and 8-strand repair methods had significantly greater strength than did the Tajima method at each time interval (p < .05 for each comparison). In addition, the 8-strand method had significantly greater normalized load values than did the Savage method at each time interval (p < .05 for each comparison). Normalized stiffness (experimental/control) for the 8-strand repair method was significantly greater than that for the Tajima and Savage methods at 3 and 6 weeks after surgery (p < .05). In addition, the normalized stiffness values for the 6-week groups was significantly greater than those for the 3-week groups (p < .05). It was concluded that the method of tendon suture was a significant variable insofar as the regaining of tendon strength was concerned and that the newer low-profile 8-strand repair method significantly expands the safety zone for the application of increased in vivo load during the early stages of rehabilitation.

Animals↗
PubMed1998

The effects of proximal load on the excursion of autogenous flexor tendon grafts.

To determine the relationship between the intrinsic properties of tendon and tendon excursion in 2 types of autogenous tendon grafts, hindpaw intrasynovial and extrasynovial tendons were transferred to the lateral and medial forepaws of adult mongrel dogs (16 experimental and 16 control tendons). After each digit was marked with radiopaque markers, it was placed in a specialized jig, and weights of 25 g, 100 g, or 200 g were applied to the flexor tendons. Specialized radiographs were obtained with the digit in flexion and extension. Tendon excursion and joint rotation were calculated. There were no statistically significant differences between experimental groups for tendon excursion or for joint angular rotation (p > .05). For intrasynovial tendon grafts, the angular rotation per millimeter tendon excursion was 10.9 degrees +/- 2.4 degrees/mm, 9.4 degrees +/- 1.2 degrees/mm, and 9.0 degrees +/- 1.4 degrees/mm with 25-g, 100-g, and 200-g loads, respectively. Comparisons between experimental groups revealed that a statistically significant difference could not be determined (p > .05). With varying proximal loads, both intrasynovial and extrasynovial grafts showed similar amounts of tendon excursion. Variations in proximal load did not significantly influence the amount of excursion within the range of loads tested. Based on these data, it appears that the difference in repair potential for these 2 types of tendon grafts is not related to a differential in the amount of tendon graft excursion following transfer to the synovial space.

Animals↗
PubMed1998

Ex vivo and in vivo adenovirus-mediated gene therapy strategies induce a systemic anti-tumor immune defence in the B16 melanoma model.

The efficacy of adenovirus-mediated gene therapy for treatment of metastatic B16 melanomas, established in syngeneic C57BL/6 mice, was assessed via an ex vivo cytokine vaccine approach or via an in vivo strategy utilizing combination cytokine/herpes simplex virus-thymidine kinase (HSV-tk) suicide gene delivery and treatment with ganciclovir (GCV). In the ex vivo tumor vaccine approach, B16 melanoma cells, transduced in vitro by adenovirus containing either interleukin (IL)-2, granulocyte-macrophage colony stimulating factor (GM-CSF), or tumor necrosis factor-alpha cytokine genes and gamma irradiated, were subcutaneously injected into the flank and a distant subcutaneous challenge injection of unmodified B16 melanoma cells was performed 15 d later. Significant reductions in challenge tumor volume were observed in the IL-2 group (75% reduction; p = 0.02) and in the GM-CSF group (88% reduction; p = 0.0006), whereas the effect for tumor necrosis factor-alpha was not statistically significant. In the in vivo treatment of established melanomas, this cytokine approach was combined with a suicide gene therapy and subcutaneous B16 melanomas were directly injected with (i) IL-2/recombinant, replication-deficient adenovirus (adv) and thymidine kinase (tk)/adv, (ii) GM-CSF/adv, IL-2/adv, and tk/adv, or (iii) control beta-galactosidase (beta-gal)/adv and tk/adv. After intraperitoneal application of GCV (10 mg per kg) for 6 d, the residual tumor masses were excised and the animals challenged with unmodified B16 cells. Challenge tumor growth was reduced by 56% for the IL-2/tk/adv/GCV treatment (p = 0.041) and by 77% for the GM-CSF/IL-2/tk/adv/GCV treatment p (p = 0.037), in comparison with the beta-gal/tk/GCV control group. These data may hold significant promise for the development of effective ex vivo and in vivo gene therapy modalities to counter the highly metastatic nature of human melanoma.

Adenoviridae↗
PubMed1998

Quantitative analysis of collagen fibrils of human cruciate and meniscofemoral ligaments.

The ultrastructural anatomy of collagen fibril diameters in the cruciate and meniscofemoral ligaments, from four young human cadaver knees (mean age, 20 years, range, 17-22 years) was studied using transmission electron microscopy. Samples were harvested from the proximal, middle, and distal regions of the anterior and posterior cruciate ligaments, and the meniscofemoral ligament. Photomicrographs were taken and assessed quantitatively using image analysis software to determine the collagen fibril diameters and eccentricities, and the percentage of total cross sectional area occupied by collagen. The collagen fibril diameter for the anterior cruciate ligament was found to be largest in the distal region but it decreased as it moved proximally. The posterior cruciate ligament had an opposite trend because it decreased from the proximal to the distal region. For the meniscofemoral ligament, the fibrils of the middle region were larger than those of the proximal and distal regions. The percentage of total cross sectional area occupied by collagen, however, did not vary significantly between regions. Fibril eccentricity also varied little between ligament or location. The variability observed in fibril diameters may account for the different mechanical properties of the ligaments.

Adolescent↗
PubMed1998

A dynamic analysis of glenohumeral motion after simulated capsulolabral injury. A cadaver model.

We used a dynamic shoulder-testing apparatus and nine fresh-frozen, entire upper extremities from cadavera to evaluate the effects of varying degrees of capsulolabral injury on the kinematics of the glenohumeral joint during abduction in the scapular plane and external rotation. Joint kinematics were recorded with use of a six-degrees-of-freedom magnetic tracking device before and after the creation of each capsulolabral lesion in a progressive manner. Dislocation did not occur after simulation of a large Bankart lesion or even after sectioning of the anterior aspect of the joint capsule. However, division of the entire joint capsule (that is, both the anterior aspect and the posterior aspect) resulted in a significant increase (p < 0.05) in posterior translation during abduction in the scapular plane, and two of the nine shoulders dislocated posteriorly. External rotation of the abducted extremity produced no increase in anterior or posterior translation.

Aged↗
PubMed1997

Combination therapy with suicide and cytokine genes for hepatic metastases of lung cancer.

Metastases of lung cancer are a major cause of treatment failure. To evaluate the therapeutic efficacy of gene therapy in metastatic lung cancer, we used adenoviral (ADV) mediated transfer of the herpes simplex virus thymidine kinase (HSV-tk) gene and the cytokine gene interleukin-2 (IL-2) to treat a murine model of metastatic lung cancer in the liver. Hepatic metastases were established by intrahepatic implantation of LL2 cells in syngeneic recipient mice. One week after tumor implantation, various replication defective ADV vectors were injected intratumorally. Treatment with a vector expressing the HSV-tk followed by ganciclovir administration with ADV.tk resulted in significant regression of tumor (p<0.01) as well as prolongation of survival (p<0.001). While a vector expressing mouse IL-2 ADV.IL-2 alone was ineffective, combination therapy with HSV-tk resulted in further tumor regression and improvement of animal survival (p<0.05). These results demonstrate that suicide and cytokine genes can be utilized in combination to treat metastatic lung cancer in vivo.

Adenoviridae↗
PubMed1997

Adenovirus-mediated herpes simplex virus thymidine kinase gene and ganciclovir therapy leads to systemic activity against spontaneous and induced metastasis in an orthotopic mouse model of prostate cancer.

It is critical to develop new therapies, such as gene therapy, which can impact on both local and metastatic prostate cancer progression. We have developed an orthotopic mouse model of metastatic prostate cancer using a cell line (RM-1) derived from the mouse prostate reconstitution (MPR) model system. This mouse model closely simulates the anatomical and biological milieu of the prostate and allows for realistic testing of experimental gene therapy protocols. Adenovirus (ADV)-mediated transduction of the herpes simplex virus thymidine kinase (HSV-tk) gene in conjunction with ganciclovir (GCV) in this model led to significant suppression of growth and of spontaneous metastasis at 14 days post-tumor inoculation. Longer-term studies produced a significant survival advantage and a continued suppression of metastatic activity for treatment animals despite regrowth of the primary tumor. Challenge by injection of tumor cells into the tail vein following excision of treated and control s.c. primary tumors resulted in 40% reduction in lung colonization in the treatment group, indicating the possible production of systemic anti-metastatic activity following a single in situ treatment with ADV/HSV-tk + GCV in this model system.

Adenoviridae↗
PubMed1997

Effect of growth factors on matrix synthesis by ligament fibroblasts.

Although it has been reported that several growth factors modulate soft-tissue healing, the specific effects of growth factors on protein synthesis during ligament healing have not been widely investigated. In this study, we examined the effects of basic and acidic fibroblast growth factors, transforming growth factor beta 1, and epidermal growth factor on collagen and noncollagenous protein synthesis by cultured fibroblasts from medial collateral ligament and anterior cruciate ligament in vitro. Uptake of tritiated proline was used to measure synthesis of collagen and noncollagenous protein, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis was used to analyze the type of collagens synthesized. Our data showed that transforming growth factor beta 1 increased both collagen and noncollagenous protein synthesis by medial collateral and anterior cruciate ligament fibroblasts on a dose-dependent basis. Collagen synthesis by cultured fibroblasts from the medial collateral and anterior cruciate ligaments was increased by treatment with transforming growth factor beta 1 by as much as approximately 1.5 times that of untreated controls. Although the response to transforming growth factor beta 1 by anterior cruciate ligament fibroblasts was equal to that by medial collateral ligament fibroblasts, the amounts of matrix proteins synthesized by anterior cruciate ligament fibroblasts were approximately half of that by medial collateral ligament fibroblasts. The increase was mostly in type-I collagen. Treatment of anterior cruciate ligament fibroblasts with epidermal growth factor increased collagen synthesis by approximately 25% but had little effect on medial collateral ligament fibroblasts. Neither basic nor acidic fibroblast growth factor increased either collagen or noncollagenous protein synthesis. These findings suggest that topical application of transforming growth factor beta 1, alone or in combination with epidermal growth factor, may have the potential to strengthen the ligament by increasing matrix synthesis during its remodeling and healing processes.

Analysis of Variance↗