A ROLE FOR BIOELECTRIC EFFECTS IN THE INDUCTION OF BYSTANDER SIGNALS BY IONIZING RADIATION?

The induction of “bystander effects” i.e. effects in cells which have not received an ionizing radiation track, is now accepted but the mechanisms are not completely clear. Bystander effects following high and low LET radiation exposure are accepted but mechanisms are still not understood. There is some evidence for a physical component to the signal. This paper tests the hypothesis that bioelectric or biomagnetic phenomena are involved. Human immortalized skin keratinocytes and primary explants of mouse bladder and fish skin, were exposed directly to ionizing radiation or treated in a variety of bystander protocols. Exposure of cells was conducted by shielding one group of flasks using lead, to reduce the dose below the threshold of 2mGy 60Cobalt gamma rays established for the bystander effect. The endpoint for the bystander effect in the reporter system used was reduction in cloning efficiency (RCE). The magnitude of the RCE was similar in shielded and unshielded flasks. When cells were placed in a Faraday cage the magnitude of the RCE was less but not eliminated. The results suggest that liquid media or cellcell contact transmission of bystander factors may be only part of the bystander mechanism. Bioelectric or bio magnetic fields may have a role to play. To test this further, cells were placed in a Magnetic Resonance Imaging (MRI) machine for 10min using a typical head scan protocol. This treatment also induced a bystander response. Apart from the obvious clinical relevance, the MRI results further suggest that bystander effects may be produced by non-ionizing exposures. It is concluded that bioelectric or magnetic effects may be involved in producing bystander signaling cascades commonly seen following ionizing radiation exposure

Multiphasic scaffold for scapholunate interosseous ligament reconstruction: a study in the rabbit knee

Scapholunate interosseous ligament tears are a common wrist injury in young and active patients that can lead to suboptimal outcomes after repair. This research aims to assess a multiphasic scaffold using 3D‐printing for reconstruction of the dorsal scapholunate interosseous ligament. The scaffold was surgically implanted in vivo in the position of the native rabbit medial collateral ligament. Two branches of treatment were implemented in the study. In the first group, the rabbits (n = 8) had the knee joint fixed in flexion for 4 weeks using 1.4 mm K‐wires prior to sample harvesting. The second group (n = 8) had the rabbit knee joint immobilized for 4 weeks prior to K‐wire removal and mobilization for an additional 4 weeks prior to sample harvesting. Overall, samples were harvested at 4 weeks post‐surgery (immobilized group) and eight weeks post‐surgery (mobilized group). Mechanical tensile testing (n = 5/group) and histology (n = 3/group) of the constructs were conducted. Tissue integration and maturation were observed resulting in increased mechanical strength of the operated joint at 8 weeks (P

Bystander Effects Induced by Exposure to Sublethal Radiation and Heavy Metals in Atlantic Salmon

These experiments were designed to identify cellular effects in 3 key organs in Atlantic Salmon (Salmo salar, L.) after exposure in vivo to very low doses of radiation, and subtoxic levels of aluminum (Al) and cadmium (Cd) or copper (Cu) alone or in combination. The salmon (approx. 35g) were exposed to doses of 4, 18 and 75 mGy doses of gamma radiation, respectively, administered over 5 hrs in untreated lake water or with Cd, Al or Cd+Al added to lake water. Six fish per group were sacrificed after exposure and the head kidney, fin and gill were dissected and sent for tissue culture. The Al and Cd speciation in the water and the levels in fish gills were measured as well as physiological parameters associated with stress. Small explants of each tissue were set up as tissue cultures using RPMI 1640 medium supplemented with serum, insulin, hydrocortisone and antibiotics. After 2 days, the culture medium was harvested and filtered then placed on a reporter cell line for determination of stress signal activity (bystander effects). Radiation doses as low as 4 mGy alone or in combination with Cd and / or Al, caused bystander signals to be produced in tissues harvested from in vivo exposed salmon. The effects vary between different organs and are not consistently additive or synergistic for a given treatment. Individual results were recorded for each individual fish. Some individual fish were consistently more sensitive to the stressors. Tissue type also appears to be critical, with gill cells showing high degrees of synergism between radiation and metal exposure. Most data for Cd suggests that lower toxicity is found when the metal is used in combination with radiation exposure. This may be due to competition between calcium and Cd since calcium is a key component in the signal transduction pathway being followed. The results indicate that this stress signal response will be a useful indicator of combined environmental stress in species inhabiting aquatic ecosystems

VEGF functionalization of suture tape results in decreased graft inflammatory and catabolic response in a rabbit model of ACL reconstruction

IntroductionImproving ligament reconstruction biology may potentially be achieved through capturing autologous circulating factors such as vascular endothelial growth factor (VEGF) using commercially available biomaterials.ObjectivesTo evaluate anterior cruciate ligament reconstruction (ACLR) using a VEGF functionalized suture tape in a rabbit model of ACLR with a semitendinosus autograft.MethodsVEGF-binding peptides were covalently bonded to polyethylene suture tape (ST) to generate functionalized constructs. Forty-six female New Zealand white rabbit ACLs were reconstructed with semitendinosus hamstring autograft (n = 6), hamstring + ST (n = 16), hamstring + scrambled peptide ST (n = 17), and hamstring + VEGF-functionalized ST (n = 17). Healing was evaluated at 2to 4 weeks using PCR, RNA sequencing, micro CT, histology, and biomechanical testing.ResultsAll rabbits successfully underwent ACLR, with no adverse events. ACLR with VEGF ST demonstrated significant decreases in inflammatory response (CD14, CD163), catabolism (MMP1, MMP3), and apoptosis (TNFSF10, Caspase-10) markers as compared to nonfunctionalized ST (P ≤ .04). µCT demonstrated similar bone tunnel mineral density in hamstring + VEGF ST rabbits as compared to hamstring + scrambled ST controls (P ≥ .31). Histology and biomechanical testing similarly demonstrated no adverse effects of VEGF-based immunomodulation on the tendon grafts.ConclusionsUsing a rabbit model, ACLR performed with VEGF-functionalized suture tape demonstrated significantly decreased markers of inflammation, catabolism, and apoptosis as compared to ACLR with nonfunctionalized suture tape. No adverse effects of functionalization were noted. Future studies should further investigate the utility of functionalized suture tape in ACLR

Fresh Osteochondral Allograft Transplantation in the Knee: A Viability and Histologic Analysis for Optimizing Graft Viability and Expanding Existing Standard Processed Graft Resources Using a Living Donor Cartilage Program

Objective: This study aims to (1) determine and validate living cartilage allograft transplantation as a novel source for viable osteochondral allograft (OCA) tissues and (2) perform histologic and viability comparisons of living donor cartilage tissues to currently available clinical-grade standard processed grafts. Design: Using healthy cartilage from well-preserved contralateral compartments in 27 patients undergoing total knee arthroplasty (TKA) and 10 clinical-grade OCA specimens obtained immediately following operative implantation, standard and living donor OCA quality was evaluated at the time of harvest and following up to 3 weeks of storage on the basis of macroscopic International Cartilage Repair Society grade, histology, and viability. Results: Osteochondral samples demonstrated a consistent decrease in viability and histologic quality over the first 3 weeks of storage at 37°C, supporting the utility of an OCA paradigm shift toward early implantation, as was the clinical standard up until recent adoption of transplantation at 14 to 35 days following donor procurement. Samples from the 10 clinical-grade OCAs, implanted at an average of 23 days following graft harvest demonstrated a mean viable cell density of 45.6% at implantation, significantly lower (P < 0.01) than the 93.6% viability observed in living donor allograft tissues. Conclusions: Osteochondral tissue viability and histologic quality progressively decreases with ex vivo storage, even when kept at physiologic temperatures. Currently available clinical OCAs are stored for 2 to 5 weeks prior to implantation and demonstrate inferior viability to that of fresh osteochondral tissues that can be made available through the use of a living donor cartilage program

A multi-chamber tissue culture device for load-dependent parallel evaluation of tendon explants

Background Injuries in the musculoskeletal system, such as tendon and ligament ruptures, are challenging to manage and often require surgical reconstructions with limited long-term success. Thus, characterizations of these tissues are urgently needed to better understand cellular mechanisms that regulate tissue homeostasis and healing. Explant culturing systems allow for ex vivo analysis of tissues in an environment that mimics the native microenvironment in vivo. Methods Collaborative efforts within our institution facilitated the establishment of a novel explant culturing system. Tissue specimens cultured in single wells, with individual applied loading and/or biological environment, allowed characterization of tissue cultured under a variety of biological loading conditions. Quantitative PCR analysis for selected gene markers was our primary outcome. Results Data were stratified for analysis by either culture environment or loading condition. Our gene expression results show that specimens clustered by culture condition may differ in molecular markers related to ECM production (e.g., Col1a1, Adamts4) and/or organization (e.g., Tnc, Dnc). In contrast, loading condition did significantly alter the median gene expression levels of tissues in comparison to unloaded control samples, although gene expression values related to ECM degradation (e.g., Mmp1, Mmp10) were altered in tendons cultured under tension in the device. Conclusion Our study demonstrates promising utility of a novel explant culturing system for further characterization of musculoskeletal tissues such as native tendons and ligaments, as well as pathologic fibrotic tissues resulting from arthrofibrosis or Dupuytren’s disease

Modernizing Storage Conditions for Fresh Osteochondral Allografts by Optimizing Viability at Physiologic Temperatures and Conditions

Objective. Osteochondral allograft (OCA) transplantation has demonstrated good long-term outcomes in treatment of cartilage defects. Viability, a key factor in clinical success, decreases with peri-implantation storage at 4°C during pathogen testing, matching logistics, and transportation. Modern, physiologic storage conditions may improve viability and enhance outcomes. Design. Osteochondral specimens from total knee arthroplasty patients (6 males, 5 females, age 56.4 ± 2.2 years) were stored in media and incubated at normoxia (21% O2) at 22°C or 37°C, and hypoxia (2% O2) at 37°C. Histology, live-dead staining, and quantitative polymerase chain reaction (qPCR) was performed 24 hours after harvest and following 7 days of incubation. Tissue architecture, cell viability, and gene expression were analyzed. Results. No significant viability or gene expression deterioration of cartilage was observed 1-week postincubation at 37°C, with or without hypoxia. Baseline viable cell density (VCD) was 94.0% ± 2.7% at day 1. At day 7, VCD was 95.1% (37°C) with normoxic storage and 92.2% (37°C) with hypoxic storage (P ≥ 0.27). Day 7 VCD (22°C) incubation was significantly lower than both the baseline and 37°C storage values (65.6%; P < 0.01). COL1A1, COL1A2, and ACAN qPCR expression was unchanged from baseline (P < 0.05) for all storage conditions at day 7, while CD163 expression, indicative of inflammatory macrophages and monocytes, was significantly lower in the 37°C groups (P < 0.01). Conclusion. Physiologic storage at 37°C demonstrates improved chondrocyte viability and metabolism, and maintained collagen expression compared with storage at 22°C. These novel findings guide development of a method to optimize short-term fresh OCA storage, which may lead to improved clinical results