The composition of cell-based therapies obtained from point-of-care devices/systems which mechanically dissociate lipoaspirate: a scoping review of the literature.

PURPOSE: Cell-based therapies using lipoaspirate are gaining popularity in orthopaedics due to their hypothesised regenerative potential. Several 'point-of-care' lipoaspirate-processing devices/systems have become available to isolate cells for therapeutic use, with published evidence reporting their clinical relevance. However, few studies have analysed the composition of their 'minimally-manipulated' cellular products in parallel, information that is vital to understand the mechanisms by which these therapies may be efficacious. This scoping review aimed to identify devices/systems using mechanical-only processing of lipoaspirate, the constituents of their cell-based therapies and where available, clinical outcomes. METHODS: PRISMA extension for scoping reviews guidelines were followed. MEDLINE, Embase and PubMed databases were systematically searched to identify relevant articles until 21st April 2022. Information relating to cellular composition and clinical outcomes for devices/systems was extracted. Further information was also obtained by individually searching the devices/systems in the PubMed database, Google search engine and contacting manufacturers. RESULTS: 2895 studies were screened and a total of 15 articles (11 = Level 5 evidence) fulfilled the inclusion criteria. 13 unique devices/systems were identified from included studies. All the studies reported cell concentration (cell number regardless of phenotype per millilitre of lipoaspirate) for their devices/systems (range 0.005-21 × 106). Ten reported cell viability (the measure of live cells- range 60-98%), 11 performed immuno-phenotypic analysis of the cell-subtypes and four investigated clinical outcomes of their cellular products. Only two studies reported all four of these parameters. CONCLUSION: When focussing on cell concentration, cell viability and MSC immuno-phenotypic analysis alone, the most effective manual devices/systems were ones using filtration and cutting/mincing. However, it was unclear whether high performance in these categories would translate to improved clinical outcomes. Due to the lack of standardisation and heterogeneity of the data, it was also not possible to draw any reliable conclusions and determine the role of these devices/systems in clinical practice at present. LEVEL OF EVIDENCE: Level V Therapeutic.

Artificial intelligence for image analysis in total hip and total knee arthroplasty : a scoping review.

AIMS: Total hip arthroplasty (THA) and total knee arthroplasty (TKA) are common orthopaedic procedures requiring postoperative radiographs to confirm implant positioning and identify complications. Artificial intelligence (AI)-based image analysis has the potential to automate this postoperative surveillance. The aim of this study was to prepare a scoping review to investigate how AI is being used in the analysis of radiographs following THA and TKA, and how accurate these tools are. METHODS: The Embase, MEDLINE, and PubMed libraries were systematically searched to identify relevant articles. The Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for scoping reviews and Arksey and O'Malley framework were followed. Study quality was assessed using a modified Methodological Index for Non-Randomized Studies tool. AI performance was reported using either the area under the curve (AUC) or accuracy. RESULTS: Of the 455 studies identified, only 12 were suitable for inclusion. Nine reported implant identification and three described predicting risk of implant failure. Of the 12, three studies compared AI performance with orthopaedic surgeons. AI-based implant identification achieved AUC 0.992 to 1, and most algorithms reported an accuracy > 90%, using 550 to 320,000 training radiographs. AI prediction of dislocation risk post-THA, determined after five-year follow-up, was satisfactory (AUC 76.67; 8,500 training radiographs). Diagnosis of hip implant loosening was good (accuracy 88.3%; 420 training radiographs) and measurement of postoperative acetabular angles was comparable to humans (mean absolute difference 1.35° to 1.39°). However, 11 of the 12 studies had several methodological limitations introducing a high risk of bias. None of the studies were externally validated. CONCLUSION: These studies show that AI is promising. While it already has the ability to analyze images with significant precision, there is currently insufficient high-level evidence to support its widespread clinical use. Further research to design robust studies that follow standard reporting guidelines should be encouraged to develop AI models that could be easily translated into real-world conditions. Cite this article: Bone Joint J 2022;104-B(8):929-937.

Combined quadriceps tendon reconstruction and total knee replacement with computer navigation: a case report.

BACKGROUND: Chronic quadriceps tendon rupture is technically challenging for surgeons due to scarring and tendon retraction. The presence of concurrent ipsilateral knee osteoarthritis compounds the issue even further. Although a combined presentation is rare, treatment options to manage each coexisting pathology simultaneously are scarcely reported. We describe the case of a 67-year-old Caucasian male who had such a presentation, and was subsequently treated with a one-stage extensor mechanism autograft reconstruction and total knee replacement with computer navigation. CASE: The patient was a 67-year-old male Caucasian, who had previously sustained an acute rupture of his right quadriceps tendon that was adequately repaired 6 months prior. Despite an initial satisfactory result, he reported deterioration in his mobility in the few months thereafter, with worsening knee pain. His comorbidities consisted of hypertension, asthma, and a body mass index of 40.4 kg/m2. Otherwise, there were no risk factors for tendon rerupture. Clinical examination later revealed a large palpable gap in the right suprapatellar region and weakness of active knee extension. No traumatic cause for this new presentation was identified. Suspicion of a chronic quadriceps tendon rupture was confirmed on radiological imaging, but the investigations also noted the presence of severe tricompartmental osteoarthritis of the ipsilateral, native knee joint. The combined procedure took place in one surgical sitting. The total knee replacement with patella resurfacing was performed first and assisted by computer navigation. The quadriceps tendon reconstruction was then conducted sequentially using the patient's hamstring tendons (semitendinosus and gracilis). The tensile strength was reinforced with use of a Ligament Augmentation and Reconstruction System (LARS) ligament. Initial outcomes were excellent, and these results were maintained at 6 months postoperatively, with the patient reporting no pain and having full range of movement. CONCLUSION: Our techniques used have not previously been reported, but are successful options in treating coexisting chronic quadriceps tendon rupture and ipsilateral knee osteoarthritis. The advantage of using computer navigation with an extramedullary femoral jig can lead to improved accuracy of bone cuts, which is important in the presence of anatomical disruption. Chronic failures of the extensor mechanism require different approaches depending on the inherent and underlying pathology. We feel that the multidisciplinary team approach to the management and use of two surgeons with differing expertise added to the successful outcome of this complex case.

Blockade of placental growth factor reduces vaso-occlusive complications in murine models of sickle cell disease.

Vaso-occlusive crisis (VOC) is the most common and debilitating complication of sickle cell disease (SCD); recurrent episodes cause organ damage and contribute to early mortality. Plasma placental growth factor (PlGF) levels are elevated in SCD and can further increase under hypoxic conditions in SCD mice. Treatment with a PlGF-neutralizing antibody (anti-PlGF Ab) in SCD mice reduced levels of monocyte chemoattractant protein-3, eotaxin, macrophage colony-stimulating factor, and plasminogen activator inhibitor-1 significantly, and of macrophage-derived chemokine and macrophage inflammatory protein-3ß moderately; this may contribute to inhibition of leukocyte recruitment, activation, and thrombosis. In subsequent experiments, anti-PlGF Ab treatment significantly reduced plasma lactate dehydrogenase levels, indicating possible reduction in cellular destruction and/or hemolysis. Histopathology studies revealed decreased incidence and severity of congestion in the lungs and spleen with repeated anti-PlGF Ab treatment. Furthermore, anti-PlGF Ab significantly reduced vaso-occlusion events under hypoxic conditions in a modified dorsal skinfold chamber model in SCD mice. Therefore, elevated PlGF levels may contribute to recruitment and activation of leukocytes. This can subsequently lead to increased pathology of affected organs in addition to mediating acute hypoxia/reoxygenation-triggered vaso-occlusion under SCD conditions. Thus, targeting PlGF may offer a therapeutic approach to reduce acute VOC and possibly alleviate long-term vascular complications in patients with SCD.

Noncovalent stabilization of the factor VIII A2 domain enhances efficacy in hemophilia A mouse vascular injury models.

An important negative regulator of factor VIIIa (FVIIIa) cofactor activity is A2 subunit dissociation. FVIII molecules with stabilized activity have been generated by elimination of charged residues at the A1-A2 and A2-A3 interfaces. These molecules exhibited reduced decay rates as part of the enzymatic factor Xa generation complex and retained their activities under thermal and chemical denaturing conditions. We describe here the potency and efficacy of 1 such stability variant, D519V/E665V, derived from B domain-deleted FVIII (BDD-FVIII). The major effect of A2 stabilization was on cofactor activity. D519V/E665V potency was increased twofold by the 2-stage chromogenic assay relative to BDD-FVIII. D519V/E665V demonstrated enhanced thrombin generation responses (fivefold by peak thrombin) relative to BDD-FVIII. In vivo consequences of enhanced cofactor activity of D519V/E665V included >fourfold increased maximal platelet-fibrin deposition after laser injury and twofold increased protection from bleeding in acute and prolonged vascular injury model in hemophilia A mice. These results demonstrate that noncovalent stabilization of the FVIII A2 subunit can prolong its cofactor activity, leading to differential enhancement in clot formation over protection from blood loss in hemophilia. The FVIII molecule described here is the first molecule with clear efficacy enhancement resulting from noncovalent stabilization of the A2 domain.

Infratemporal fossa reconstruction following total auriculectomy: an alternative flap option.

Reconstruction following oncologic resection in the head and neck is complex due to large surgical defects left after removal of skin, subcutaneous, and skeletal structures. It is essential to adequately fill the defect as well as provide an acceptable tissue match in terms of tone, texture, thickness and contour. A 55-year-old male presented with an advanced melanoma in the right pre-tragal area. Surgical resection was performed including a total auriculectomy. A tunnelled right supraclavicular artery island (SAI) flap was used to repair the surgical defect. A Doppler probe ensured adequate circulation within the flap, especially in the distal tip. Reconstruction using the SAI flap after oncologic ear resection reduced operating room time, required less technical expertise, and provided excellent tissue match compared to more traditional methods of surgical defect reconstruction including free flaps, local flaps, and pedicled myocutaneous flaps. Successful use of the SAI flap in this case further expands the flaps versatility. We recommend that the reconstructive surgeon consider the SAI flap when presented with challenging infratemporal fossa and lateral skull base cases.

Supraclavicular artery island flap for head and neck oncologic reconstruction: indications, complications, and outcomes.

BACKGROUND: The supraclavicular island flap has been used successfully for difficult facial reconstruction cases, providing acceptable results without using microsurgical techniques. The authors use this regional flap in reconstructing various head and neck oncologic defects that normally require traditional regional or free flaps to repair surgical wounds. METHODS: A pedicled supraclavicular artery flap was used to reconstruct head/neck oncologic defects. Complications and functional outcomes were assessed. RESULTS: Head and neck oncologic patients underwent tumor resection followed by immediate reconstruction using a supraclavicular artery island flap. Ablative defects included neck, tracheal-stomal, mandible, parotid, and pharyngeal walls. All flaps (n = 18) were harvested in less than 1 hour. All ablative wounds and donor sites were closed primarily and did not require additional surgery. Major complications included a complete flap loss when the vascular pedicle was inadvertently divided and pharyngeal leaks. The leaks resolved without surgical intervention, and both patients regained the ability to swallow using their neo-esophagus. Minor complications included donor-site wound dehiscence and cellulitis. None of the patients reported functional donor-site morbidity. CONCLUSIONS: This thin flap is easy and quick to harvest, has a reliable pedicle, and has minimal donor-site morbidity. It is now the authors' flap of choice for many common head and neck reconstructive problems. Early experience using the supraclavicular artery island flap suggests that it is an excellent flap option for head and neck oncologic disease patients.

Supraclavicular artery flap: a new option for pharyngeal reconstruction.

Laryngopharyngeal oncologic resections produce complex reconstructive problems, requiring dependable robust flaps to restore form and function. Current options include morbid local-regional flaps or free tissue transfers. The supraclavicular artery flap (SAF) offers a great new option. Partial pharyngeal oncologic defects were reconstructed with pedicled SAFs. Handheld Doppler probes marked the pedicle preoperatively. Flaps were design based upon the dopplered vascular anatomy. Complications and functional outcomes were assessed. All flaps (n = 6) were harvested in under 1 hour with uneventful postoperative recoveries. Ablative wounds and donor sites were closed primarily. Two patients had small controlled leaks because of preoperative radiation and overly aggressive oral intakes, that subsequently resolved. There were no functional donor site morbidities. We describe a novel application of the SAF for pharyngeal reconstructions after laryngopharyngeal cancer ablation. This thin, reliable, easy to harvest, low morbidity flap is an excellent reconstructive option for pharyngeal reconstructions.

Hair follicle melanocyte cells as a renewable source of melanocytes for culture and transplantation.

OBJECTIVE: Advances in melanocyte culture techniques have not yet led to reliable clinical methods for treating hypopigmentation disorders. We hypothesized that melanocytes harvested from plucked hair follicles may provide a renewable source of melanocytes for the treatment of hypopigmentation. METHODS: Hairs with attached cells from the follicles were plucked from Yucatan pigs and implanted in a collagen-glycosaminoglycan matrix for either immediate or delayed implantation into full-thickness excisional porcine wounds. Wounds were allowed to heal and were biopsied at 2 and 4 weeks, respectively. RESULTS: Fully healed wounds with transplanted hair follicles showed central areas of dark pigmentation corresponding to the location of implanted hair follicles. Corresponding collagen-glycosaminoglycan matrix wounds showed no central areas of pigmentation. CONCLUSIONS: Hair follicle--derived melanocytes may potentially serve as a renewable source of pigment-producing cells for treating hypopigmentation disorders.

Tensile forces stimulate vascular remodeling and epidermal cell proliferation in living skin.

OBJECTIVES: To quantify tissue remodeling induced by static and cyclical application of tensional forces in a living perfused tissue. BACKGROUND: Cells are able to respond to mechanical cues from the environment and can switch between proliferation and quiescence. However, the effects of different regimens of tension on living, perfused skin have not been characterized. METHODS: The ears of living rats were mechanically loaded by applying tensile forces (0.5 Newtons) either statically or cyclically and then analyzing tissue responses using in vivo microscopy, immunohistochemistry, and corrosion casting. RESULTS: Quantitative immunohistochemistry showed that in the static group (4-day continuous tension) there was up to 4-fold increase in cellular proliferation in the epidermis after 4 days and a 2.8-fold increase in the vascularity in the dermis that peaked after 2 days. Comparable effects could be achieved in just 8 hours using a cyclic loading protocol. We also modeled the resultant stress produced in the ear using a linear finite element model and demonstrated a correlation between the level of applied stress and both epidermal cell proliferation and blood vessel density. CONCLUSIONS: Mechanical forces stimulate cell proliferation and vascular remodeling in living skin. As cell growth and vascular supply are critical to wound healing and tissue expansion, devices applying controlled mechanical loads to tissues may be a powerful therapy to treat tissue defects.

Side population hematopoietic stem cells promote wound healing in diabetic mice.

BACKGROUND: Early evidence suggests that stem cells play a role in normal wound healing. Various impaired wound-healing states might be due to a deficiency in the stem cell repertoire. The authors sought to demonstrate that a new subset of lymphoid progenitor murine hematopoietic stem cells will accelerate wound healing in diabetic mice. METHODS: Bone marrow cells were harvested from C57Bl6/J femurs and separated into side and main populations based on their ability to efflux the vital dye Hoechst 33342 and the presence or absence of CD7 and CD34 markers. Side or main population cells and control solution were applied once topically to 1-cm full-thickness dorsal excisional wounds in lepr db/db and wild-type mice on the day after wounding (n = 12 in each group). Wound closure was followed by computer planimetry. Wounds were harvested after 7 and 25 days for histological analysis. RESULTS: Topical side population treatment had a significant effect on wound closure in diabetic animals, with a higher percentage of wound closure (35 +/- 7.2 percent) in this group on postoperative day 7 compared with animals treated with either main population cells (16 +/- 4.9 percent) or a vehicle control using saline (14 +/- 6.7 percent) (p < 0.05). When side population cells were given to wild-type mice that already had a normal stem cell repertoire, there was a trend toward better wound closure, but no significant differences were found. CONCLUSIONS: Side population-treated wounds healed more quickly than main population-or control-treated wounds in diabetic mice, suggesting that one stem cell subpopulation, but not the majority, harbors the potential for improving the healing process. Further studies are needed to investigate the mechanism of healing and to explore its potential as a therapeutic agent.

Optimization of UV cross-linking density for durable and nontoxic collagen GAG dermal substitute.

Artificial dermal constructs, based upon collagen-glycosaminoglycan matrices (CGMs), provide new options in treating skin defects. However, their clinical effectiveness may be limited by cytotoxicity related to residual aldehydes left over from the manufacturing process. Although both chemical and dehydrothermal (DHT) cross-linking are used to produce CGMs, we hypothesize that optimized nonchemical cross-linking, using ultra-violet (UV) and DHT treatment combinations, may limit cytotoxicity without sacrificing mechanical strength. Porous CGMs were physically cross-linked using a combination of DHT and varying intensities of UV light. These were compared to glutaraldehyde cross-linked controls. Human keratinocytes were seeded in each matrix, and cellular proliferation measured using a microculture tetrazolium dye assay. A scoring system (based on the in vitro contraction rate, stiffness, and cellular growth of a small cylindrical specimen) was developed to assess the best overall physical cross-linking method. More cellular growth was observed in the 90-120 min UV cross-linked group than in the glutaraldehyde-treated group (p < 0.05). Stiffness was maximized after 0-30 min of UV cross-linking. On the basis of our scoring system, DHT combined with 45 min of UV cross-linking produced the best overall matrix in terms of cellular growth and physical durability. UV cross-linked collagen-based biomaterials could be a viable alternative for use in biological applications to eliminate glutaraldehyde-associated cytotoxicity.

Gene-based delivery of IFN-beta is efficacious in a murine model of experimental allergic encephalomyelitis.

Experimental allergic encephalomyelitis (EAE) is a model of central nervous system (CNS) inflammation that follows immunization with certain CNS antigens. The course and clinical manifestations of EAE are similar to those of multiple sclerosis (MS) in humans; therefore, EAE has become an accepted animal model to study MS. The purpose of this study was to demonstrate that systemic expression of murine interferon-beta (IFN-beta) (MuIFN-beta), following intramuscular (i.m.) delivery of plasmid DNA encoding MuIFN-beta to the hind limb of mice, is effective in reducing the clinical manifestations of disease in a model of EAE. The results of the study demonstrate that gene-based delivery of MuIFN-beta caused significantly decreased clinical scores compared with delivery of the null vector. A single injection of the MuIFN-beta plasmid was as effective in reducing the severity of the disease as an every other day injection of MuIFN-beta protein.

Age-dependent acceleration of ischemic injury in endothelial nitric oxide synthase-deficient mice: potential role of impaired VEGF receptor 2 expression.

Morbidity and mortality of peripheral arterial occlusive disease significantly increases with age, often exhibiting more severe disease pathology and decreased treatment effectiveness. Therapeutic angiogenesis with angiogenic growth factors may represent a valuable treatment option for the severely ill, older adult patient population. Aging is considered an independent cardiovascular risk factor, but pathomechanistically it is not well understood. Diminished endothelial nitric oxide (EDNO) production has been considered as a major contributor to the aging process. To investigate the effect of age on postischemic revascularization independent of changes in EDNO, we used endothelial nitric oxide synthase-deficient (ecNOS-KO) mice. We found an age-dependent acceleration in ischemic injury following unilateral femoral artery ligation in these animals compared to C57BL/J6 mice. Postischemic revascularization, quantified by measuring von Willebrand factor expression, was significantly impaired, suggesting that factors other than progressive EDNO deterioration are also involved in the age-dependent severe disease phenotype. Ischemia led to an increase in the expression of vascular endothelial growth factor receptor-2, KDR, in younger ecNOS-KO; however, this increase in KDR expression was absent in the older animals. Lack of increased KDR expression may provide a mechanistic explanation for the severe ischemic injury and perhaps can be used as a clinical marker to identify severe, vascular endothelial growth factor refractory patient population.

Effect of recombinant platelet-derived growth factor (Regranex) on wound closure in genetically diabetic mice.

Burns, especially those involving large surface areas, represent a complex wound healing problem. Platelet-derived growth factor (PDGF) is released by activated platelets to recruit inflammatory cells toward the wound bed. It has effects on promoting angiogenesis and granulation tissue formation. However, the effectiveness of topical PDGF on wound closure is variable, ranging from little improvement observed in pig models to dramatic improvement reported in a diabetic mouse model. Here, we sought to determine the effectiveness of commercially sold PDGF-BB (Regranex) on wound closure in genetically diabetic mice. C57BL/KsJ db+/db+ mice and its host strain bearing dorsal 1.5-cm wounds were divided into groups (n = 8 in each group) receiving topical application of either Regranex (10 microg/wound) or vehicle for 5 consecutive days after wounding. The rate of wound closure was analyzed using computerized planimetry. The amount of granulation tissue was determined histologically. Our data indicate that diabetic mice exhibit a significant delay in wound closure when compared with their host strain. Topical application of Regranex did not improve the time to wound closure but did significantly increase the amount of granulation tissue. Our current study using commercially available Regranex failed to reproduce the previously reported finding that PDGF improved wound closure in healing impaired genetically diabetic mice.

Expired liquid preserved platelet releasates retain proliferative activity.

BACKGROUND: Donated platelets for clinical use currently have a shelf life of 5 days as the result of possible bacterial contamination and loss of hemostatic function. Platelet releasates contain multiple growth factors that have been shown to accelerate wound healing. We sought to demonstrate that although expired platelets can no longer sustain hemostasis, they serve a longer term role as a reservoir of growth factors that could be harnessed in wound healing applications. MATERIALS AND METHODS: Liquid preserved human platelets were activated from 1 to 21 days after collection using zeolite and were then analyzed for their ability to stimulate human fibroblast proliferation, which is an in vitro serogate of growth factor activity and wound healing potential. Total protein content, the concentration of platelet-derived growth factor (PDGF) and transforming growth factor-beta were also measured. RESULTS: Activated liquid preserved platelet releasates significantly stimulated fibroblast proliferation. Twenty-one-day-old platelets were as stimulatory as 2-day-old platelets. Total protein concentration, PDGF, and transforming growth factor-beta concentrations remained constant throughout the 21-day course. Western blot analysis using an antibody against human PDGF revealed minimal protein degradation over time. CONCLUSIONS: These data demonstrate that although the role of platelets as hemostatic agents degrades rapidly with time, platelets' ability to serve as a reservoir for growth factors remains intact for at least 3 weeks. These growth factors could be collected, stored, and used as a topical agent to promote healing of chronic and recalcitrant wounds.