Biomedical subjects

H Uludag

Publications and source records attributed to H Uludag.

22 records · Page 2Linked to original sources

PubMedDate not supplied

Bone affinity of a bisphosphonate-conjugated protein in vivo.

Growth factors capable of stimulating bone formation are potential therapeutic agents for osteoporosis treatment. It is essential, however, that a targeting mechanism is incorporated into the growth factors to deposit them at osseous tissue with minimal distribution to extraskeletal sites. To this end, a strategy has been developed in which a bone-seeking molecule, 1-amino-1,1-diphosphonate methane (aminoBP), was chemically conjugated to a model protein, bovine serum albumin (BSA). This study was carried out to assess the bone affinity of the conjugates in a tibia injection model. Using ovariectomized (OVX) rats, initial (3 h) retention of BSA and aminoBP-BSA were found to be equivalent when injected into the medullary cavity of tibia. After 1 day, an 8- and 12-fold higher tibiae retention of the protein was obtained in normal and OVX rats as a result of aminoBP conjugation. A similar result ( approximately 12-fold difference) was also obtained in OVX rats after 3 days. We concluded that aminoBP conjugation to BSA imparted a high bone affinity and enhanced bone retention of proteins in normal and OVX rats.

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PubMedDate not supplied

Bisphosphonate conjugation to proteins as a means to impart bone affinity.

Growth factors are endogenous proteins capable of stimulating new bone formation, but their clinical benefit for systemic stimulation of bone mass has not been demonstrated. The critical challenge is to deliver a significant dose of the proteins to bone after intravenous injection. This challenge may be overcome by derivatizing proteins with ligands that exhibit a high bone affinity (e.g., bisphosphonates). To demonstrate the feasibility of this approach, 1-amino-1,1-diphosphonate methane (aminoBP) was conjugated to a model protein, albumin. The conjugation was performed by (1) converting the amino group of aminoBP to a thiol group using 2-iminothiolane, (2) derivatizing the albumin amino groups with a thiol-reactive sulfosuccinimidyl-4-(N-maleimidomethyl)-1-cyclohexane carboxylate, and (3) reacting the derivatized albumin with thiolated aminoBP. Typically, 1-4 aminoBP molecules per albumin were obtained. The conjugated albumin exhibited a high affinity to hydroxyapatite that was proportional to the extent of conjugation. The conjugates were shown to exhibit a high affinity to bone matrix in vitro in a serum-containing medium. Once bound to bone matrix, the conjugates were found to desorb more slowly than the unmodified albumin, especially from bone whose organic matrix was removed by ashing. In conclusion, conjugation of bisphosphonates to albumin was shown to impart a high bone affinity to the protein, and such conjugates can be potentially targeted to bone.

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PubMedDate not supplied

Control of water flux in a bioartificial kidney.

UC-PK1 cells were grown to confluence on microporous microcellulose membranes in order to test the feasibility of using transmembrane pressure (TMP) for controlling connective water transport in a bioartificial kidney. TMP was applied on either the apical or basal aspect of the polarized cell layer, while the fluid compartments on both sides of the membrane were perfused with tissue culture medium in a miniature flow chamber. The cell monolayer did not allow filtration in the apical to basal direction when positive TMP up to 30 mmHg was applied on the apical side. Application of positive TMP on the basal side led to measurable ultrafiltration. The hydraulic permeability, L(p), of the cell-seeded membranes was found to be increasing with time, reaching a steady state value after 60 min. There was a strong positive correlation between L(p) and applied TMP. For a constant TMP, L(p) was found to be independent of the shear rate between 2.6 and 10.5 sec-1. When positive pressure was changed abruptly back from the basal to apical site, the ultrafiltration rate decreased to zero within seconds. Morphologic studies suggest that the tight junctions between cells were broken by TMP applied on the basolateral aspect of the cell monolayer. It is concluded that, basolateral TMP may be used to control water flux in a bioartificial kidney.

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PubMedDate not supplied

Metabolic activity and proliferation of CHO cells in hydroxyethyl methacrylate-methyl methacrylate (HEMA-MMA) microcapsules.

To better understand encapsulated cell behaviour, Chinese Hamster Ovary (CHO) fibroblasts were encapsulated in HEMA-MMA microcapsules and short-term (<2 wks) proliferation and changes in metabolic activity were investigated in vitro. CHO cells were observed to undergo rapid proliferation in the first week following encapsulation after which a growth arrest was obtained at approximately 3500 cells/capsule. The cell growth was localized in aggregates in the capsule core, resulting in high local cell density but low cell density in the whole capsule interior (approximately 10(7) cells/mL). The total metabolic activity, as determined by the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium) assay, within the microcapsules increased in the first week, with no significant change afterwards. A broad variation in metabolic activity among the individual capsules was obtained. Supplementing the cell suspension with 20% Ficoll 400 during the encapsulation process resulted in significantly higher morphological uniformity among the individual capsules (with reduced capsule wall thickness and eccentricity); however, this did not change the extent of heterogeneity in metabolic activity. We conclude that viability and proliferation ability (at least to a limited extent) of CHO cells are maintained in HEMA-MMA microcapsules. The local cell growth and subsequent growth arrest remain issues to be addressed in order to obtain better utilization of the microcapsule core volume. Alternatively, small diameter (< 400 microns as opposed to the present approximately 750 microns diameter) capsules are necessary.

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