Mrinal Musib

Synergistic Rate Boosting of Collagen Fibrillogenesis in Heterogeneous Mixtures of Crowding Agents

The competition for access to space that arises between macromolecules is the basis of the macromolecular crowding phenomenon, known to modulate biochemical reactions in subtle ways. Crowding is a highly conserved physiological condition in and around cells in metazoans, and originates from a mixture of heterogeneous biomolecules. Here, using collagen fibrillogenesis as an experimental test platform and ideas from the theory of nonideal solutions, we show that an entropy-based synergy is created by a mixture of two different populations of artificial crowders, providing small crowders with extra volume occupancy when in the vicinity of bigger crowders. We present the physiological mechanism by which synergistic effects maximize volume exclusion with the minimum amount of heterogeneous crowders, demonstrating how the evolutionarily optimized crowded conditions found in vivo can be reproduced effectively in vitro.

Artificial intelligence in research

We asked young scientists to describe an example of artificial intelligence or machine learning in research, its broader implications in the field, and the challenges scientists face when using such technologies. Our surveys responses reflected a variety of countries and fields, but only 6% came

MICROHARDNESS OF BI-ANTIBIOTIC-ELUTING BONE CEMENT SCAFFOLDS

Bi-antibiotic-impregnated bone cements (BIBCs) are widely used in orthopaedics as a prophylactic agent (depot) to address post-surgical infections. Although hardness is widely considered a viable index to measure the integrity of the cement structure, there are few specific studies involving changes in hardness characteristics of BIBCs post elution of high doses of two widely used antibiotics: tobramycin and gentamicin. Increased doses of antibiotics and increased duration of elution may also decrease the hardness of polymethyl methacrylate (PMMA) bone cement, thus increasing the chances of shattering, scratching, and deformation.In this project, we have investigated the changes in surface hardness of five different antibiotic-loaded specimens: 0.5 g tobramycin and 0.5 g gentamicin together, 1 g tobramycin, 1 g gentamicin, 5 g tobramycin and 5 g gentamicin together, and 10 g tobramycin (each added to 40 g of PMMA), post elution for various time periods (1, 3, and 21 days). The effect of hydration on the hardness of bone cement was studied to replicate in vivo conditions. The micro-indentation tester (Buehler m5103) was utilized to determine if the increased antibiotic loads would compromise the integrity of the bone cement matrix.The results demonstrated that the amount of drug initially incorporated determined the hardness of the cement post elution. As compared to the control (no antibiotic), specimens containing 1 and 10 g of antibiotic exhibited over 50% and 73% decrease in hardness, respectively. The different treatment durations (post 1 day) as well as the hydration conditions had insignificant effect on the hardness of the cement.

Biomechanical Evaluation of Osteoporotic Sheep Long Bones

The objective of this research was to develop a calcium phosphate-based synthetic bone mineral (SBM) that will have a potential for osteoporosis therapy and prevention. The goal of this study was to test the efficacy of SBM in preventing loss of strength in long bones of osteoporotic sheep. The animals were divided into four groups: sham-operated; ovariectomized (OVX); OVX supplemented with SBM without fluoride; OVX supplemented with SBM containing fluoride. The bones were scanned by CT and tested mechanically in 4-point-bending from which the mechanical properties and fracture behavior were evaluated. The load at 1mm deformation for the OVX sheep tibias was significantly lower than the sham operated group and groups with special diets. The sheep given the SBM (+F) showed the highest load carrying capacity. Similarly, the stiffness and energy to fracture of the OVX sheep was also significantly lower than the group given SBM (+F). The ultimate stress and the elastic modulus did not show any statistically significant difference among the four groups. SBM was partially successful at preventing loss of bone strength in osteoporotic sheep.

Phagocytosis of Simulated Nano-wear Debris by Osteoblasts

In this study I have used well characterized simulated wear-nano-particles of ultra-high molecular weight polyethylene (UHMWPE) of known size and shape to study particulate phagocytosis by MG63 osteoblast-like cells. The particles were treated to decrease their propensity to form aggregates in aqueous suspension and scanning electron microscopy (SEM) was performed to image them as individual particles on 0.01 µm pore size polycarbonate filter membranes. These images were further subjected to morphometric analysis of the particles using ASTM F1877 descriptors (equivalent circle diameter (ECD in μm), aspect ratio (AR), elongation (E), roundness (R), and formfactor (FF)). The mean (±SD) ECD of the particles were 0.056±0.03 µm. They were subsequently introduced to confluent MG63 cells at particle:cell ratio of 100:1 and incubated for 24 h. Transmission electron micrography (TEM) showed the nanoparticles inside the cytosol. No time dependent response was observed beyond 24 h. The nanoparticles seemed to re-agglomerate once inside the cell. The damage to the cells was evident from the compromised cell membrane. This study will help further our knowledge of the wear-mediated osteolytic process.

Effect of wear-debris particles on RAW 264.7 cells

Wear-debris particles from ultrahigh molecular weight polyethylene (UHMWPE) and metals has been implicated in periprosthetic osteolysis and cellular response to them depend on their size and dose. Response to larger particles has been previously studied, but there are no specific studies examining biological response to simultaneous administration of both UHMWPE and Ti particles less than 0.2μm in size. Our lab has developed a standardized and reproducible technique to isolate, characterize and fractionate both particles into 3 distinct size ranges [1.0–10.0μm (micron), 0.2–1.0μm (submicron) and 0.01–0.2μm (nano)] and furthermore minimize clumping, which facilitated disaggregation and isolation of individual particles. RAW 264.7 cells were treated for 24h, 48h and 72h with various doses and sizes of both particulates simultaneously. Cell membrane damage was measured by quantifying lactate dehydrogenase (LDH) release. The cells elicited a dose and size dependent response to the nano-particulates at the highest dose. At 72h there was a significant increase in LDH released by cells treated with the nanoparticles as compared to untreated controls as well as to larger particles. To mimic in vivo conditions, further studies are being conducted and will involve administration of dual wear particulates that will help understand cellular response to wear particulates.

Nano-Wear-Particulates Elicit a Size and Dose Dependent Response by RAW 264.7 Cells

Ultrahigh molecular weight polyethylene (UHMWPE) and metals, primarily Ti and Co-Cr alloys are widely used to make components of orthopaedic implants. Cellular response to their wear-particulates depends on their size and dose. Although biological response to larger particles has been previously studied, there are no specific studies examining cellular response to simultaneous administration of well characterized clinically relevant size ranges of both UHMWPE and Ti particles less than 0.2 μm in size. The overall hypothesis for this work is that RAW 264.7 cells will display a more robust negative response to nanaoparticulates as compared to larger particles. Isolation of ’clean’ simulated wear particles has been an issue and we in our lab have been able to develop a standardized and reproducible technique to isolate, characterize and fractionate (following ASTM F-1877 standards) both UHMWPE and Ti particles into 3 distinct size ranges [1.0-10.0 μm (micron), 0.2-1.0 μm (submicron) and 0.01-0.2 μm (nano)] from periprosthetic tissue explants and furthermore minimize clumping, thus facilitate disaggregation and isolation of individual particles into micron-, submicron and nano-fractions. Cells were treated for 24 h, 48 h and 72 h with various doses (103, 105, 107) and sizes (mentioned above) of both UHMWPE and Ti particles. Proliferation was determined by counting the number of cells at the specified time periods. Preliminary studies reveal early stimulatory and late inhibitory effect on cell proliferation. At 24 h, the nanofraction particles elicited a proliferative response by the cells, though statistically not significant, but at 72 h there was a significant inhibitory response. The response was dose and size dependent and the cells exhibited an inhibitory response to the nanofraction particulates only at the highest dose. To mimic in vivo conditions, further studies are being conducted and will involve administration of dual wear particulates that will help understand cellular response to wear particulates.