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Biomedical subjects

David A Lee

Publications and source records attributed to David A Lee.

At least 19 recordsLinked to original sources
PubMed2006

The influence of swelling and matrix degradation on the microstructural integrity of tendon.

Tendon is multi-level fibre composite material, responsible for the transmission of forces from muscles to the skeleton. It is composed of a hierarchical arrangement of collagenous units surrounded by a proteoglycan-rich matrix, arranged to support strain transfer, and thus contribute to the mechanical behaviour of tendon. This study examines the effect of swelling and enzymatic degradation on structural integrity at different levels of the tendon hierarchy. Biochemical and microstructural analysis are used to examine the effects of incubation on the composition and swelling of the matrix, prior to a mechanical characterisation of sample integrity. Results indicated significant swelling of tendon fibrils and interfibrillar matrix after incubation in phosphate buffered saline, leading to a reduction in ultimate tensile load, with failure initiated between fibrils and sub-fibrils. In contrast, incubation with the enzyme chondroitinase ABC resulted in a total removal of glycosaminoglycan from the samples, and a subsequent reduction in the extent of swelling. These fascicles also demonstrated an increase in failure loads, with failure predominating between fibres. The findings from this work confirm the importance of the non-collagenous matrix components in controlling strain transfer within tendon structures. It also highlights the necessity to maintain samples within a suitable and controlled environment prior to testing.

Animals↗
PubMed2006

Are all melanomas the same? Spitzoid melanoma is a distinct subtype of melanoma.

BACKGROUND: Although the majority of melanomas demonstrate high rates of mutations in B-RAF or N-RAS that result in constitutive activation of the mitogen-activated protein kinase-signaling pathway, emerging data suggest molecular differences among melanoma subtypes. In this study, the authors evaluated the contribution of B-RAF and N-RAS mutations to the pathogenesis of Spitzoid melanomas. METHODS: In total, 33 Spitzoid melanomas were analyzed for clinical and pathologic characteristics as well as for hot-spot mutations in the B-RAF and N-RAS genes. In the majority of patients (28 of 33 melanomas), the tumors were confined to the skin with no evidence of metastasis (average follow-up, 32.5 mos). There were five metastasizing melanomas (5 of 33 tumors) with regional or systemic spread. RESULTS: Of 33 Spitzoid melanomas, only 1 showed the V600E mutation in the B-RAF gene (1 of 33 tumors; 3%). It was noteworthy that none of the metastatic Spitzoid melanomas (0 of 5 tumors; 0%), of which 2 resulted in fatal outcomes, demonstrated mutations in B-RAF or N-RAS. CONCLUSIONS: In contrast to the majority of cutaneous melanomas, activating hot-spot mutations in B-RAF or N-RAS were not involved in the pathogenesis of Spitzoid melanoma. These data suggested that Spitzoid melanoma is a distinct form of melanoma with unknown genes and/or signaling pathways involved in its development.

Adolescent↗
PubMed2006

Rate of oxygen consumption by isolated articular chondrocytes is sensitive to medium glucose concentration.

Cartilage tissue engineering typically involves the culture of isolated chondrocytes within a scaffold material. The oxygen tension within the engineered tissue is known to be an essential parameter for implant success. This will be sensitive to the oxygen consumption behavior of the embedded chondrocytes, which remains to be characterized. We report that the oxygen consumption of bovine articular chondrocytes is sensitive to glucose deprivation below 2.7 mM, increasing from a basal level of 9.6x10(-16) to <18.4x10(-16) mol/cell.h in 1.3 mM glucose. Further studies examined the influence of selecting high (18.4 mM) or low (5.1 mM) glucose medium on the oxygen tension in 2 mm thick cellular agarose constructs. A relative upregulation of oxygen consumption was observed in constructs cultured in low glucose medium. This resulted in the near-anoxic oxygen concentration of 5 microM oxygen in constructs seeded with 40x10(6) cells/ml, compared to 57 microM in the corresponding high glucose culture. The upregulation of oxygen consumption generally corresponded to the inhibition of glycolysis, which is consistent with the Crabtree phenomenon. Medium osmolarity (316-600 mOsm) had minimal effects on chondrocyte oxygen consumption rate. In conclusion, glucose availability is a critical parameter that regulates the oxygen tension within tissue engineered constructs.

Animals↗
PubMed2006

Economic impact of preoperative curettage before Mohs micrographic surgery for basal cell carcinoma.

BACKGROUND: The role of curettage before Mohs micrographic surgery for basal cell carcinoma (BCC) remains controversial. Preoperative curettage may allow the surgeon to better delineate the subclinical extensions of high-risk BCCs, thereby enabling a more precise first-stage excision around tumor-containing tissue. OBJECTIVE: To assess the economic impact of preoperative curettage for high-risk BCCs treated with Mohs micrographic surgery on patients, providers, and insurers. METHODS: Given the enormous variability in practice styles, it was estimated that the time required to complete a second stage of Mohs surgery was 25, 50, or 75% of that required to complete the first stage. New York City Medicare and Standard reimbursement rates were used to approximate the cost of an additional stage of Mohs surgery for high-risk BCCs. RESULTS: Assuming that preoperative curettage increases operative efficiency by reducing the number of required Mohs stages from 2 to 1, the time saved can be quantified. Thus, if the Mohs surgeon estimates that the time required to remove a second stage is 75% of that of the first stage, the time savings with preoperative curettage equals 75% of the duration of a one-stage Mohs surgery. Similarly, when a second stage requires 50 or 25% of the time needed to complete the first stage, the time saved equals 50 or 25% of the duration of a one-stage Mohs surgery. Reducing the number of stages from 2 to 1 saves insurers and privately paying patients approximately $250 and $500, respectively. CONCLUSIONS: Whether preoperative curettage can offer a more precise first-stage excision without compromising tissue conservation remains a subject of debate. Preoperative curettage may reduce the number of Mohs surgical stages required for tumor clearance, potentially shortening patient encounters and allowing surgeons to treat additional patients, while decreasing costs for patients and insurers.

Basal Cell Carcinoma↗
PubMed2005

Cyclic tensile strain upregulates collagen synthesis in isolated tendon fascicles.

Mechanical stimulation has been implicated as an important regulatory factor in tendon homeostasis. In this study, a custom-designed tensile loading system was used to apply controlled mechanical stimulation to isolated tendon fascicles, in order to examine the effects of 5% cyclic tensile strain at 1 Hz on cell proliferation and matrix synthesis. Sample viability and gross structural composition were maintained over a 24 h loading period. Data demonstrated no statistically significant differences in cell proliferation or glycosaminoglycan production, however, collagen synthesis was upregulated with the application of cyclic tensile strain over the 24 h period. Moreover, a greater proportion of the newly synthesised matrix was retained within the sample after loading. These data provide evidence of altered anabolic activity within tendon in response to mechanical stimuli, and suggest the importance of cyclic tensile loading for the maintenance of the collagen hierarchy within tendon.

Adaptation, Physiological↗
PubMed2005

Glaucoma and its treatment: a review.

PURPOSE: A review of glaucoma and its treatment is presented. SUMMARY: Glaucoma is a common eye disease that can cause irreversible blindness if left undiagnosed and untreated. Glaucoma is a leading cause of blindness in the United States and other industrialized countries. In most cases, the symptoms of early-stage glaucoma are minimal or nonexistent. There are several different types of glaucoma, and they have been classically divided into the categories of primary or secondary open-angle or angle-closure glaucoma. Secondary forms of glaucoma are caused by various ocular or systemic diseases. Every available treatment to prevent progressive glaucomatous optic neuropathy has potential adverse effects and involves a certain amount of risk and financial expense. Conventional first-line treatment of glaucoma usually begins with the use of a topical selective or nonselective beta-blocker or a topical prostaglandin analog. Second-line drugs of choice include alpha-agonists and topical carbonic anhydrase inhibitors. Parasympathomimetic agents, most commonly pilocarpine, are considered third-line treatment options. For patients who do not respond to antiglaucoma medications, laser trabeculoplasty and incisional surgery are further methods that can be used to lower intraocular pressure. The results of clinical trials have reaffirmed the utility of antiglaucoma medications in slowing the progression of the disease. CONCLUSION: Glaucoma is a common eye disease that is usually associated with an elevated intraocular pressure. Treatment options for patients with glaucoma include medications, laser therapy, and incisional surgery. The risks and benefits of each type of treatment must be carefully considered to maximize the treatment's benefits while minimizing adverse effects.

Administration, Topical↗
PubMed2005

The influence of noncollagenous matrix components on the micromechanical environment of tendon fascicles.

Tendon is composed of type I collagen fibers, interspersed with proteoglycan matrix and cells. Glycosaminoglycans may play a role in maintaining the structural integrity of tendon, preventing excessive shearing between collagen components. This study tests the hypothesis that tendon extension mechanisms can be altered by modifying the composition of noncollagenous matrix. Tendon explants were treated with phosphate buffered saline (PBS) or PBS + 0.5 U ml(-1) chondroitinase ABC. Structural changes were examined using TEM and biochemical analysis, while strain response was examined using confocal microscopy and gross mechanical characterization. Chondroitinase ABC removed 90% of glycosaminoglycans from the matrix. Results demonstrated significant swelling of fibrils and surrounding matrix when incubated in either solution. In response to applied strain, PBS incubated samples demonstrated significantly less sliding between adjacent fibers than nonincubated, and a 33% reduction in maximum force. By contrast, fascicles incubated in chondroitinase ABC demonstrated a similar strain response to nonincubated. Data indicate that collagen-proteoglycan binding characteristics can be influenced by incubation and this, in turn, can influence the preferred extension mechanisms adopted by fascicles. This highlights the importance of maintaining fascicles within their natural environment to prevent structural or mechanical changes prior to subsequent analysis.

Animals↗
PubMed2005

Ocular hypotensive medications for the treatment of glaucoma.

The past 10 years have seen dramatic changes in the pharmacologic treatment of glaucoma. The treatment of glaucoma has changed from surgical to primarily medical. These treatments have improved the effectiveness and safety of lowering the intraocular pressure to prevent progressive vision loss and blindness, but no cure for glaucoma or reversal of blindness is available yet. Research is progressing and can hopefully achieve these final goals sometime in the future.

Antihypertensive Agents↗
PubMed2005

Mutations in the CYLD gene in Brooke-Spiegler syndrome, familial cylindromatosis, and multiple familial trichoepithelioma: lack of genotype-phenotype correlation.

Brooke-Spiegler syndrome (BSS), familial cylindromatosis (FC), and multiple familial trichoepithelioma (MFT), originally described as distinct entities, share overlapping clinical findings. Patients with BSS are predisposed to multiple skin appendage tumors such as cylindroma, trichoepithelioma, and spiradenoma. FC, however, is characterized by cylindromas and MFT by trichoepitheliomas as the only tumor type. These disorders have recently been associated with mutations in the CYLD gene. In this report, we describe three families with BSS, one with FC, and two with MFT phenotypes associated with novel and recurrent mutations in CYLD. We provide evidence that these disorders represent phenotypic variation of a single entity and lack genotype-phenotype correlation.

Carcinoma, Adenoid Cystic↗
PubMed2005

Nutrient utilization by bovine articular chondrocytes: a combined experimental and theoretical approach.

A combined experimental-numerical approach was adopted to characterize glucose and oxygen uptake and lactate production by bovine articular chondrocytes in a model system. For a wide range of cell concentrations, cells in agarose were supplemented with either low or high glucose medium. During an initial culture phase of 48 h, oxygen was monitored noninvasively using a biosensor system. Glucose and lactate were determined by medium sampling. In order to quantify glucose and oxygen uptake, a finite element approach was adopted to describe diffusion and uptake in the experimental model. Numerical predictions of lactate, based on simple relations for cell metabolism, were found to agree well for low glucose, but not for high glucose medium. Oxygen did not play a role in either case. Given the close association between chondrocyte energy metabolism and matrix synthesis, a quantifiable prediction of utilization can present a valuable contribution in the optimization of tissue engineering conditions.

Animals↗
PubMed2004

Activin receptor-like kinase-7 induces apoptosis through activation of MAPKs in a Smad3-dependent mechanism in hepatoma cells.

Activin receptor-like kinase (ALK)7 is a type I serine/threonine kinase receptor of the transforming growth factor (TGF)-beta family of proteins that has similar properties to other type I receptors when activated. To see whether ALK7 can induce apoptosis as can some of the other ALK proteins, we infected the FaO rat hepatoma cell line with adenovirus expressing a constitutively active form of the ALK7. Cells infected with active ALK7 adenovirus showed an apoptotic-positive phenotype, as opposed to those that were infected with a control protein. DNA fragmentation assays and fluorescence-activated cell sorter analysis also indicated that ALK7 infection induced apoptosis in FaO cells. We also confirmed this finding in Hep3B human hepatoma cells by transiently transfecting the constitutively active form of ALK7, ALK7(T194D). Investigation into the downstream targets and mechanisms involved in ALK7-induced apoptosis revealed that the TGF-beta signaling intermediates, Smad2 and -3, were activated, as well as the MAPKs JNK and p38. In addition, caspase-3 and -9 were also activated, and cytochrome c release from the mitochondria was observed. Short interfering RNA-mediated inhibition of Smad3 markedly suppressed ALK7-induced caspase-3 activation. Treatment with protein synthesis inhibitors or the expression of the dominant-negative form of the stress-activated protein/extracellular signal-regulated kinase 1 abolished not only JNK activation but apoptosis as well. Taken together, these results suggest that ALK7 induces apoptosis through activation of the traditional TGF-beta pathway components, thus resulting in new gene transcription and JNK and p38 activation that initiates cross-talk with the cellular stress death pathway and ultimately leads to apoptosis.

Activin Receptors, Type I↗
PubMed2004

EyeSite: a semi-automated database of protein families in the eye.

The EyeSite is a web-based database of protein families for proteins that function in the eye and their homologous sequences. The resource clusters proteins at different levels of homology in order to facilitate functional annotation of sequences and modelling of proteins from structural homologues. Eye proteins are organized into the tissue types in which they function and are clustered into homologous families using a novel protocol employing the TribeMCL algorithm. Homologous families are further subdivided into sequence clusters for which multiple sequence alignments are generated. Structural annotations from the CATH domain database are provided for nearly 90% of the sequences, and protein family annotations from the Pfam database for approximately 86%. Homology models have also been generated where appropriate. The EyeSite is stored in a relational database and is extensively linked to other online bioinformatics resources to help relate allelic variants, annotations and clinical details to the derived data in the database. The EyeSite is available for online search, sequence information and model retrieval at http://eyesite.cryst.bbk.ac.uk/.

Amino Acid Sequence↗
PubMed2004

Crosslinking density influences the morphology of chondrocytes photoencapsulated in PEG hydrogels during the application of compressive strain.

Chondrocyte deformation, which occurs during mechanical loading, is thought to play an important role in the mechanotransduction pathway. In designing a scaffold that can be gelled in situ for cartilage tissue engineering, an important consideration is the influence of mechanical loading. This study tested the hypothesis that changes in the crosslinking density of a hydrogel scaffold influence the morphology of encapsulated chondrocytes in response to an applied load. Chondrocytes were entrapped in photo-crosslinkable hydrogel scaffolds based on poly(ethylene glycol) (PEG) with two crosslinking densities, 0.119 and 0.376 mol/l, with the higher density having a 11-fold higher compressive modulus. The cell-embedded hydrogels were subjected to static compressive strains between 0% and 20% after 1 and 6 days of culture. Using confocal laser scanning microscopy, chondrocytes in the highly crosslinked gel at day 1 deformed more than gels in the more loosely crosslinked gel. By day 6, this finding was reversed. When single cells within a region were followed, heterogeneities in cell deformation were observed on both a macroscopic and microscopic scale. These heterogeneities were greater in the highly crosslinked gel. These findings demonstrate that different levels of cell deformation and heterogeneity may be obtained by varying the crosslinking density in PEG hydrogels.

Animals↗
PubMed2004

Crosslinking density influences chondrocyte metabolism in dynamically loaded photocrosslinked poly(ethylene glycol) hydrogels.

In approaches to tissue engineer articular cartilage, an important consideration for in situ forming cell carriers is the impact of mechanical loading on the cell composite structure and function. Photopolymerized hydrogel scaffolds based on poly(ethylene glycol) (PEG) may be synthesized with a range of crosslinking densities and corresponding macroscopic properties. This study tests the hypothesis that changes in the hydrogel crosslinking density influences the metabolic response of encapsulated chondrocytes to an applied load. PEG hydrogels were formulated with two crosslinking densities that resulted in gel compressive moduli ranging from 60 to 670 kPa. When chondrocytes were encapsulated in these PEG gels, an increase in crosslinking density resulted in an inhibition in cell proliferation and proteoglycan synthesis. Moreover, when the gels were dynamically loaded for 48 h in unconfined compression with compressive strains oscillating from 0 to 15% at a frequency of 1 Hz, cell proliferation and proteoglycan synthesis were affected in a crosslinking-density-dependent manner. Cell proliferation was inhibited in both crosslinked gels, but was greater in the highly crosslinked gel. In contrast, dynamic loading did not influence proteoglycan synthesis in the loosely crosslinked gel, but a marked decrease in proteoglycan production was observed in the highly crosslinked gel. In summary, changes in PEG hydrogel properties greatly affect how chondrocytes respond to an applied dynamic load.

Animals↗
PubMed2004

Comparison of the safety and efficacy of dorzolamide 2% and brimonidine 0.2% in patients with glaucoma or ocular hypertension.

PURPOSE: To compare the intraocular pressure (IOP) reduction between dorzolamide 2% and brimonidine 0.2% in primary open-angle glaucoma (POAG) or ocular hypertension (OHT). METHODS: This study was a prospective, double-masked, randomized, crossover comparison of dorzolamide 2% (Trusopt) and brimonidine 0.2% (Alphagan), three times daily during two six-week study periods. The primary endpoint was mean change from baseline in trough IOP and secondary endpoints were mean change from baseline in IOP one and three hours after dosing. T-tests and a repeated-measures ANOVA were used to statistically evaluate the data. RESULTS: Of 43 patients enrolled, 41 completed the first treatment and 38 completed both treatments. Baseline IOP for dorzolamide was 24.3 mm Hg and brimonidine, 24.6 mm Hg (P = 0.9). Mean IOP reduction at trough was similar for both agents, 3.0 mm Hg (P = 0.96). Reductions at one and three hours were comparable (P = ns). Both agents were well tolerated with adverse events consistent with the package inserts. Dorzolamide was associated with more frequent stinging (P = 0.017) and burning (P < 0.001), while brimonidine was associated with more frequent dry eye (P = 0.04). CONCLUSIONS: Dorzolamide and brimonidine, as monotherapy, produced equivalent IOP-lowering efficacy at trough and at one and three hours after instillation, and both were well tolerated.

Adrenergic alpha-Agonists↗
PubMed2004

Delusional discourse: an investigation comparing the spontaneous causal attributions of paranoid and non-paranoid individuals.

Research into the nature of attributional reasoning in paranoia has for the most part been restricted to questionnaire-based approaches. This fails to address the issue of whether a distinctive attributional style underpins the everyday talk of paranoid individuals. This study aimed to investigate whether attributional models of paranoid delusions applied to spontaneous attributions generated in the discourse of 12 paranoid and 12 non-paranoid speakers. Causal attributions for negative and positive life experiences were extracted from interview transcripts and rated using the Content Analysis of Verbatim Explanations (CAVE) technique. It was found that, as a proportion, paranoids made more attributions for negative events that were of an external-personal, stable and global nature (as attributional models would predict). They also made significantly more external-personal attributions for negative events and, in one of two datasets, showed a more external mean CAVE rating for negative events than the non-paranoid controls. This paper highlights important issues underlying the extraction of attributions from paranoid talk, and discusses the implications for attributional models of paranoia and future discourse-based research in this area.

Adult↗
PubMed2004

Mechanical loading of chondrocytes embedded in 3D constructs: in vitro methods for assessment of morphological and metabolic response to compressive strain.

Mechanical loading of chondrocytes in 3D constructs has been used to investigate mechanotransduction and its potential for stimulating tissue-engineered cartilage repair. This chapter describes the preparation of 3D agarose or alginate constructs seeded with isolated chondrocytes and specific test rigs for applying gross compressive strain to individual constructs on a confocal microscope or for longer term compression of constructs cultured within an incubator. Experimental methods are described to quantify the level of cell deformation and the elaboration of extracellular matrix. The chapter thus provides an introduction to the experimental techniques used to examine chondrocyte mechanotransduction and downstream cell function.

Alginates↗