The C. elegans Taste Receptor Homolog LITE-1 Is a Photoreceptor.

Many animal tissues/cells are photosensitive, yet only two types of photoreceptors (i.e., opsins and cryptochromes) have been discovered in metazoans. The question arises as to whether unknown types of photoreceptors exist in the animal kingdom. LITE-1, a seven-transmembrane gustatory receptor (GR) homolog, mediates UV-light-induced avoidance behavior in C. elegans. However, it is not known whether LITE-1 functions as a chemoreceptor or photoreceptor. Here, we show that LITE-1 directly absorbs both UVA and UVB light with an extinction coefficient 10-100 times that of opsins and cryptochromes, indicating that LITE-1 is highly efficient in capturing photons. Unlike typical photoreceptors employing a prosthetic chromophore to capture photons, LITE-1 strictly depends on its protein conformation for photon absorption. We have further identified two tryptophan residues critical for LITE-1 function. Interestingly, unlike GPCRs, LITE-1 adopts a reversed membrane topology. Thus, LITE-1, a taste receptor homolog, represents a distinct type of photoreceptor in the animal kingdom.

Pharmacogenetics as a predictor chemotherapy induced peripheral neuropathy in gynecologic cancer patients treated with Taxane-based chemotherapy.

BACKGROUND: This study investigated whether there are pharmacogenomic markers predictive of chemotherapy induced peripheral neuropathy (CIPN) as a result of taxane-based chemotherapy. METHODS: Patients were enrolled from August 2020 to November 2020 in a prospective, case-control trial evaluating pharmacogenetic predictors of CIPN. All women were treated with at least 3 cycles of taxane-based chemotherapy for histologically confirmed gynecologic malignancies. Buccal saliva samples were used to test for 32 drug metabolism variations. All testing was performed by ⍺LPHA-GENOMIX laboratories. Fisher's Exact test was used to assess for event differences of categorical variables. RESULTS: Of 102 enrolled patients, 58%, 28%, and 14% had ovarian, endometrial, or cervical cancers, respectively. The median age was 67, 72% were Caucasian and 25% were African American. 16% of patients were treated with 3-4 cycles, 57% received 5-7 cycles, and 27% received 8 or more cycles of chemotherapy that included paclitaxel. Grade 2 CIPN was experienced by 51 patients. There was no difference in age, race, disease site, or number of chemotherapy cycles (p > 0.05) between those who did or did not develop CIPN. CYP2D6 genotype (p = 0.009) and CYP3A5 genotype (p = 0.023) had different frequencies among those with and without CIPN. Patients classified as having poor or intermediate function of CYP2D6 had increased risk of CIPN (OR 1.63, 95% CI 1.04-2.57, p = 0.026). There was no difference in CYP2D6 phenotype by race (p = 0.29). No patients with normal function of CYP3A5 developed CIPN. There were no Caucasians with normal function of CYP3A5, but 28% of African Americans had normal CYP3A5 function (p < 0.001). CONCLUSIONS: Pharmacogenomics appear associated with the development of CIPN and may be able to help personalize treatment decision making.

Transsynaptic Fish-lips signaling prevents misconnections between nonsynaptic partner olfactory neurons.

Our understanding of the mechanisms of neural circuit assembly is far from complete. Identification of wiring molecules with novel mechanisms of action will provide insights into how complex and heterogeneous neural circuits assemble during development. In the Drosophila olfactory system, 50 classes of olfactory receptor neurons (ORNs) make precise synaptic connections with 50 classes of partner projection neurons (PNs). Here, we performed an RNA interference screen for cell surface molecules and identified the leucine-rich repeat-containing transmembrane protein known as Fish-lips (Fili) as a novel wiring molecule in the assembly of the Drosophila olfactory circuit. Fili contributes to the precise axon and dendrite targeting of a small subset of ORN and PN classes, respectively. Cell-type-specific expression and genetic analyses suggest that Fili sends a transsynaptic repulsive signal to neurites of nonpartner classes that prevents their targeting to inappropriate glomeruli in the antennal lobe.

Validation of automated complex head and neck treatment planning with pencil beam scanning proton therapy.

BACKGROUND: Pencil beam scanning (PBS) proton therapy offers dosimetric advantages for several treatment sites, including head and neck (H&N). However, to achieve the optimal target coverage and robustness, these plans can be complex and time consuming to develop and optimize. Automating the treatment planning process can ensure a high-quality and standardized plan, reduce burden to the planner, and decrease time-to-treatment. We utilized in-house scripting to automate a four-field multi-field optimization (MFO) H&N planning technique. METHODS AND MATERIALS: Ten bilateral H&N patients were planned in RayStation v6 with a four-field modified-X beam configuration using MFO planning. Automation included creation of avoidance structures to control spot placement and development of standardized beams, PBS spot settings, robust optimization objectives, and patient-specific predicted planning constraints. Each patient was planned both with and without automation to evaluate differences in planning time, perceived effort and plan quality, plan robustness, and OAR sparing. RESULTS: On average, scripted plans required 3.2 h, compared to 4.3 h without the script. There was no difference in target coverage or plan robustness with or without automation. Automation significantly reduced mean dose to the oral cavity, parotids, esophagus, trachea, and larynx. Perceived effort was scaled from 1 (minimum effort) to 100 (maximum effort), and automation reduced perceived effort by 42% (p < 0.05). Two non-scripted plans required re-planning due to errors. CONCLUSIONS: Automation of this multi-beam, the MFO proton planning process reduced planning time and improved OAR sparing compared to the same planning process without scripting. Scripting generation of complex structures and planning objectives reduced burden on the planner. With most current treatment planning software, this automation is simple to implement and can standardize quality of care across all treatment planners.

The acute management of ankle fractures (Augment) study: A prospective trainee led national collaborative audit of the Boast 12 guidelines.

BACKGROUND: Ankle fractures are one of the most common fractures in adults aged 20-65 years. The British Orthopaedic Association (BOA) and British Orthopaedic Foot and Ankle Society (BOFAS) jointly produced Standards for Trauma (BOAST) BOAST 12, with the aim of reducing morbidity by standardising care of these injuries. The primary aim of the AUGMENT study was to determine the extent and clinical effect of variation from BOAST 12. METHODS: AUGMENT was a multi-centre prospective trainee led audit of consecutive patients presenting with an ankle fracture within a four-week period. Data were collected on patient demographics, comorbidities, management and 12-week outcome. The BOAST 12 standards were divided into four subgroups; documentation, imaging, management and follow-up. Percentage compliance with each subgroup was analysed. A multivariate logistic regression analysis was used to determine impact of overall compliance on likelihood of discharge in follow-up period. FINDINGS: 971 patients were included across 52 sites. The overall rate of BOAST 12 compliance was 41.7%. Variations in practice were observed in clinical documentation, especially of neurovascular status, (40.7%) and VTE assessment (61.5%). Patient management compliance with all 16 of the BOAST 12 standards was associated with a higher rate of discharge during the 12-week follow-up period (p = 0.005). CONCLUSION: AUGMENT has demonstrated that the management of ankle fractures is variable across the UK. Over half of patients had aspects of their care that were not BOAST 12 compliant. When compliance was observed, it was associated with earlier discharge from orthopaedic care.

The Current UK Consensus on the Management of Weber B and Posterior Ankle Fractures: A Questionnaire Study as Part of the Acute Management of Ankle Fractures (AUGMENT) Audit.

Ankle fractures are common conditions which are associated with significant morbidity when managed incorrectly. With the incidence due to triple by 2030, standards of practice were created by the British Orthopaedic Association and the British Orthopaedic Foot and Ankle Society to ensure optimal treatment. In spite of this, anecdotally there is still a variation in practice and therefore a questionnaire study was designed to explore clinician decision-making around Weber B and Posterior Malleolus ankle fractures. Five scenarios explored management regarding minimally and grossly displaced injuries, as well as the use of further imaging. The questionnaires were distributed via AUGMENT collaborators at their sites and at the British Orthopaedic Foot and Ankle Society congress 2018. About 315 questionnaires were completed and included in analysis. For Weber B injuries, overall there was a consensus across all respondents with minimally displaced and grossly displaced fractures being treated conservatively and operatively respectively. For Posterior Malleolus injuries, there was variation in practice between Foot and Ankle specialists and their non-Foot and Ankle colleagues. Computed tomography (CT) was more likely to be used to assess these injuries by specialists (97.50 vs 69.79%) and these injuries were more likely to be treated operatively across the board. This study identified key variation in practice of the management of Posterior Malleolar ankle fractures, including the use of imaging to further define the anatomy and the decision to operate. Foot and Ankle surgeons were more likely to organize CT scans and to surgically manage these injures.

A case report of acute hypothermia during initial inpatient clozapine titration with review of current literature on clozapine-induced temperature dysregulations.

BACKGROUND: Here we describe a unique case of clozapine-associated hypothermia during initial titration of this medication in an acute inpatient psychiatry setting. Only a handful of cases on this topic have been published. We discuss possible pharmacologic mechanisms supporting or refuting the propensity of clozapine to induce hypothermia, as well as risk factors for clozapine-induced hypothermia, and a comparison to clozapine-induced hyperthermia. CASE PRESENTATION: A 70 year-old African American female with treatment-refractory schizoaffective disorder developed hypothermia with a nadir temperature of 89 °F (31.7 °C) after 7 days on clozapine, on a total dose of 50 mg twice daily. Accompanying symptoms included bradycardia, hypotension, QTc prolongation, tachypnea, hypoxemia, and an absence of shivering. The patient was transferred to the ICU, and rewarmed within 10 h with the discontinuation of her clozapine, ziprasidone, and carvedilol. Broad spectrum antibiotics were initiated, but discontinued shortly after, as the patient had no leukocytosis, and blood cultures were negative. DISCUSSION: While hypoglycemia, hypothyroidism, sepsis, and stroke were effectively ruled out, alternative drug-disease (including chronic kidney disease), and drug-drug interactions were considered possible contributing features. Benzodiazepines, valproic acid, ziprasidone, and the numerous antihypertensive agents the patient was taking were considered as either primary or compounding factors for hypothermia. After exclusion or inclusion of these alternative causes, we calculated a score of 4 (possible) for clozapine-induced hypothermia on the Naranjo Scale. CONCLUSIONS: Clozapine-induced hypothermia may occur more commonly than clinicians believe. Practitioners should be cognizant of this potentially fatal phenomenon, and monitor for temperature dysregulations while on clozapine, especially during initial titration, in those with multiple comorbid factors, and on additional medications that may contribute to hypothermia.

Saphenous vein graft disease, pathophysiology, prevention, and treatment. A review of the literature.

BACKGROUND: The saphenous vein remains the most frequently used conduit for coronary artery bypass grafting, despite reported unsatisfactory long-term patency rates. Understanding the pathophysiology of vein graft failure and attempting to improve its longevity has been a significant area of research for more than three decades. This article aims to review the current understanding of the pathophysiology and potential new intervention strategies. METHODS: A search of three databases: MEDLINE, Web of Science, and Cochrane Library, was undertaken for the terms "pathophysiology," "prevention," and "treatment" plus the term "vein graft failure." RESULTS: Saphenous graft failure is commonly the consequence of four different pathophysiological mechanisms, early acute thrombosis, vascular inflammation, intimal hyperplasia, and late accelerated atherosclerosis. Different methods have been proposed to inhibit or attenuate these pathological processes including modified surgical technique, topical pretreatment, external graft support, and postoperative pharmacological interventions. Once graft failure occurs, the available treatments are either surgical reintervention, angioplasty, or conservative medical management reserved for patients not eligible for either procedure. CONCLUSION: Despite the extensive amount of research performed, the pathophysiology of saphenous vein graft is still not completely understood. Surgical and pharmacological interventions have improved early patency and different strategies for prevention seem to offer some hope in improving long-term patency.

Functional Outcomes Following Operative and Nonoperative Management of Weber C Ankle Fractures: A Systematic Review.

Weber C ankle fractures are unstable ankle fractures occurring above the syndesmosis. These fractures are often managed operatively, although a small population of patients are still selected for nonoperative management. This study primarily aimed to summarize the current evidence on functional outcomes for Weber C patients managed operatively and nonoperatively. Evidence on secondary outcomes such as complications and radiographic outcomes were also reviewed. This systematic search was conducted according to PRISMA guidelines. A literature search was conducted using the EMBASE, Medline, and Central databases. A total of 26 studies were included in the final analysis. All papers studied the management of Weber C fractures using open reduction and internal fixation (ORIF). Three main functional outcome scores were identified: American Orthopedic Foot and Ankle Society score, Olerud-Molander Ankle Score, and Foot and Ankle Outcome Score. Only 1 study compared operative and conservative management, which showed similar outcomes for either option (median Olerud-Molander Ankle Score 95 [range 20 to 95] vs 100 [70 to 100], respectively). Complications associated with operative management included infection, wound dehiscence, implant failure, and malunion or nonunion. The mean rate of syndesmosis malreduction was 18.2%. This study showed that operative management, regardless of the method of ORIF used, as well as nonoperative management resulted in good functional outcomes, indicating that patient selection for either method is important. However, there is limited evidence on the complications and radiographic outcomes associated with nonoperative management. We recommend further studies to compare all ORIF methods with conservative management and examine the complications associated with nonoperative management.

Surgical vs non-surgical management of Weber B fractures: A systematic review.

BACKGROUND: The aim of this systematic review was to compare surgical and non-surgical management of Weber B ankle fractures. METHODS: A systematic computer-based search was conducted using the MEDLINE (via OvidSP), EMBASE (via OvidSP) and Central databases. Data were extracted regarding functional outcome, radiological union, range of motion (RoM), infection rate and quality of life (QoL). RESULTS: There were no significant differences identified between surgical and non-surgical management of Weber B fractures with respect to functional outcome. There is a higher rate of complication following surgical management, including infection, reoperation, thromboembolic events and death. With respect to QoL and ankle RoM, this review identified no differences between surgical and non-surgical management. CONCLUSIONS: There is a need for further published literature evaluating the most efficacious management as there is a poverty of high-level research available. Currently, the available literature does not overwhelmingly favour a particular approach to Weber B ankle fractures.

RATeS (Re-Admissions in Trauma and Orthopaedic Surgery): a prospective regional service evaluation of complications and readmissions.

INTRODUCTION: All the surgeries carry risks, which may lead to readmission at a later date. At present, there is limited Trauma and Orthopaedic (T&O) specific data in the literature. As a result, a prospective regional service evaluation aimed to discover the current complication and readmission rates across all T&O procedures and identify any factors associated with these outcomes. METHODS: Data were collected at six sites across Yorkshire and Humber for all T&O procedures during October 2016. Patient demographics and procedure-specific data were collected. Post-operative complications and length of stay were recorded. All the patients were then followed up for 30 days post-discharge to determine if they experienced complications which resulted in readmission and further surgical intervention. RESULTS: 1411 patients having a total of 64 operations were recorded with 1391 completing follow-up (98.5%). Overall in-patient complication rate was 8.4% with the readmission rate being 4.4%. An ASA grade of three or more was found to be associated with readmission. Procedure-related factors such as the use of VTE prophylaxis and prophylactic antibiotics, as well as the elective nature of certain operations were negatively associated with readmission. The largest subgroup of patients was those undergoing total hip (THR) or knee replacements (TKR). For these 234 patients, the readmission rate for TKR and THR being 3.77% and 3.13%, respectively. CONCLUSIONS: This large, multi-centre project describes readmission rates following trauma and orthopaedic surgery. In the presented study, the elective nature of the procedure was associated with a reduced risk of readmission.

Minimizing the Risk of Surgical Site Infection Following Hip Fracture Operation.

Introduction: Hip fractures are the most common serious injury in the elderly, associated with disability, morbidity, and mortality. Surgical site infection (SSI) is a serious post-operative complication. This prospective cohort study outlines how our center made cumulative improvements in SSI incidence rates, reaching a 12-month average of 0.5%. Methods: All patients undergoing hip fracture operation between 2016 and 2021 were included. The primary outcome measure was confirmed SSI, according to the Public Health England definition. Results were compared with the baseline recordings by an independent SSI team in 2013. Demographic data were compared with National Hip Fracture Database records. Peri-operative infection control and wound management tactics introduced between 2014 and 2021 were collated to gain an overview care bundle. Results: Baseline recordings identified a 9.0% SSI rate in a three-month observation period. In our study, 3,138 hip fracture operative cases were completed between October 2016 and December 2021. There were 9 superficial and 32 deep infections identified, yielding an overall infection rate of 1.3%. However, when analyzing the 12-month average, there was consistent decline in SSI from the baseline 9.0% in 2013 to 0.5% in 2021 (p < 0.05). A peri-operative care bundle included pre-operative bleeding risk assessment. Intra-operatively, double preparation and draping is used for arthroplasty. Broad-spectrum antibiotic agents and tranexamic acid are administered. Meticulous hemostasis and watertight wound closure are observed. Anti-coagulated patients received negative pressure dressings. Post-operatively, a dedicated senior lead team provided daily inpatient review of patients, with urgent consultant review of all wound healing concerns. Conclusion: Patients with a hip fracture have numerous risk factors for SSI. A dedicated multi-focal tactic, adopted by a multi-disciplinary department, can yield substantial risk reduction. Each intervention is evidence based and contributes to cumulative improvement. By prioritizing infection prevention, we have minimized the need for complex infection management interventions and achieved an annual saving of £860,000 for our trust.

A panel of phenotypically and genotypically diverse bioluminescent:fluorescent Trypanosoma cruzi strains as a resource for Chagas disease research.

Chagas disease is caused by Trypanosoma cruzi, a protozoan parasite that displays considerable genetic diversity. Infections result in a range of pathological outcomes, and different strains can exhibit a wide spectrum of anti-parasitic drug tolerance. The genetic determinants of infectivity, virulence and therapeutic susceptibility remain largely unknown. As experimental tools to address these issues, we have generated a panel of bioluminescent:fluorescent parasite strains that cover the diversity of the T. cruzi species. These reporters allow spatio-temporal infection dynamics in murine models to be monitored in a non-invasive manner by in vivo imaging, provide a capability to detect rare infection foci at single-cell resolution, and represent a valuable resource for investigating virulence and host:parasite interactions at a mechanistic level. Importantly, these parasite reporter strains can also contribute to the Chagas disease drug screening cascade by ensuring that candidate compounds have pan-species in vivo activity prior to being advanced into clinical testing. The parasite strains described in this paper are available on request.

A robust and flexible baculovirus-insect cell system for AAV vector production with improved yield, capsid ratios and potency.

Manufacturing of adeno-associated viruses (AAV) for gene and cell therapy applications has increased significantly and spurred development of improved mammalian and insect cell-based production systems. We developed a baculovirus-based insect cell production system-the SGMO Helper-with a novel gene architecture and greater flexibility to modulate the expression level and content of individual Rep and Cap proteins. In addition, we incorporated modifications to the AAV6 capsid sequence that improves yield, capsid integrity, and potency. Production of recombinant AAV 6 (rAAV6) using the SGMO Helper had improved yields compared to the Bac-RepCap helper from the Kotin lab. SGMO Helper-derived rAAV6 is resistant to a previously described proteolytic cleavage unique to baculovirus-insect cell production systems and has improved capsid ratios and potency, in vitro and in vivo, compared with rAAV6 produced using Bac-RepCap. Next-generation sequencing sequence analysis demonstrated that the SGMO Helper is stable over six serial passages and rAAV6 capsids contain comparable amounts of non-vector genome DNA as rAAV6 produced using Bac-RepCap. AAV production using the SGMO Helper is scalable using bioreactors and has improved yield, capsid ratio, and in vitro potency. Our studies demonstrate that the SGMO Helper is an improved platform for AAV manufacturing to enable delivery of cutting-edge gene and cell therapies.

Cardiometabolic consequences of therapy for chronic schizophrenia using second-generation antipsychotic agents in a medicaid population: clinical and economic evaluation.

OBJECTIVE: We assessed the potential clinical and economic impact of coronary heart disease (CHD) and diabetes arising after the use of second-generation ("atypical") antipsychotic agents for the treatment of chronic schizophrenia. We compared the use of these medications in patients with a higher risk of cardiometabolic adverse events (in a higher-risk scenario) and in patients with a lower risk (in a lower-risk scenario). Our U.S.-based analysis estimated the costs of CHD and diabetes arising from antipsychotic medication-related cardiometabolic effects. METHODS: We constructed a health economic model to predict the 5-year incidence of CHD and diabetes and associated costs after treatment. In this cost-consequence model, we used CHD risk functions derived from the Framingham Heart Study and diabetes risk functions derived from the Atherosclerosis Risk in Communities (ARIC) study. Patient characteristics and treatment effects on cardiometabolic risk factors were estimated from the Clinical Trials of Antipsychotic Treatment Effectiveness (CATIE) study. We evaluated two cost-consequence scenarios: the incidence of CHD and diabetes predicted for 1,000 patients with chronic schizophrenia in a higher-risk scenario based on data from CATIE associated with olanzapine (Zyprexa) and in a lower-risk scenario with ziprasidone (Geodon). We evaluated rates of adverse outcomes for each scenario and the cost of treatment for CHD and diabetes. All costs were reported in 2011 U.S. dollars. Because Medicaid is often the payer for patients with chronic schizophrenia, all costs in this analysis were derived from the perspective of Medicaid. RESULTS: Over a period of 5 years in 1,000 patients with chronic schizophrenia, the higher-risk scenario with olanzapine showed a 9% increased incidence of CHD and a 59% increased incidence of diabetes, compared with no change in treatment from baseline. By contrast, the lower-risk scenario with ziprasidone showed a 9% reduced incidence of CHD and a 10% reduced incidence of diabetes. The higher-risk scenario led to increased CHD-related costs of $83,206 and to increased diabetes-related costs of $456,399. CONCLUSION: Our study underscores the importance of monitoring the established risk factors for CHD and diabetes in patients using second-generation antipsychotic drugs. Lower-risk agents from this class may lead to substantially decreased costs in the management of CHD and diabetes when compared with higher-risk agents.

Prevalence of apolipoprotein E4 genotype and homozygotes (APOE e4/4) among patients diagnosed with Alzheimer's disease: a systematic review and meta-analysis.

BACKGROUND: Population allele frequencies of apolipoprotein E (APOE) vary by geographic region. The purpose of this study is to summarize and evaluate published estimates for the prevalence of APOE e4 carrier status among the population diagnosed with Alzheimer's disease (AD) by geographic region and country. METHODS: A systematic review of English-language publications from January 1, 1985, through May 31, 2010, was conducted. Studies reporting APOE e4 status for patients diagnosed with AD were included in the analysis; trials and autopsies were excluded. APOE e4 data were pooled, and prevalence and 95% confidence intervals (CIs) were calculated. RESULTS: Pooled estimates for APOE e4 carrier prevalence data were derived from 142 independent samples: 48.7% (95% CI: 46.5-51.0), and from 73 samples for e4/4 (homozygotes): 9.6% (95% CI: 8.4-10.8). The highest estimates were in Northern Europe: 61.3% (95% CI: 55.9-66.7), e4/4 prevalence: 14.1% (95% CI: 12.2-16.0). The lowest estimates were in Asia and Southern Europe. Substantial heterogeneity of these prevalence estimates was observed. CONCLUSIONS: APOE e4 genotype prevalence varies among AD patients by region and within each country. Further exploration is warranted to better understand the substantial heterogeneity of these prevalence estimates.

Mild cognitive impairment: disparity of incidence and prevalence estimates.

BACKGROUND: The purpose of conducting this study was to identify areas of concordance and sources of variation for the published rates of prevalence and incidence associated with various definitions for mild cognitive impairment (MCI). METHODS: The study used systematic review of studies published in English since 1984. Studies were identified by searching MEDLINE and EMBASE databases. Population-based observational studies of incidence or prevalence of MCI and related terms were eligible for inclusion. RESULTS: A total of 3,705 citations were identified, and 42 were accepted for inclusion; 35 included data on prevalence and 13 on incidence. The following four terms predominated: age-associated memory impairment (AAMI); cognitive impairment no dementia (CIND); MCI; and amnestic MCI (aMCI). Within each term, the operational definition varied. Substantial variation was observed for both incidence (MCI: 21.5-71.3; aMCI: 8.5-25.9 per 1,000 person-years) and prevalence of each definition of cognitive impairment (AAMI 3.6%-38.4%; CIND 5.1%-35.9%; MCI 3%-42%; aMCI 0.5%-31.9%). CIND and MCI showed increasing prevalence among older age groups, whereas age-specific rates of aMCI were lower and without any apparent age relationship. CONCLUSIONS: Prevalence and incidence estimates associated with MCI vary greatly both between definitions and within a definition across the 42 publications. These wide differences pose a significant challenge to our understanding of the social burden of this disease. Enhancement and standardization of operational definitions of the subtypes of cognitive impairment could improve estimates of disease burden and provide a mechanism to assist in the identification of individuals at risk for future Alzheimer's disease and other dementias.

Transient expression of myofibroblast-like cells in rat rib fracture callus.

BACKGROUND AND PURPOSE: We have previously shown that early fracture callus of rat rib has viscoelastic and contractile properties resembling those of smooth muscle. The cells responsible for this contractility have been hypothesized to be myofibroblast-like in nature. In soft-tissue healing, force generated by contraction of myofibroblasts promotes healing. Accordingly, we tried to identify myofibroblast-like cells in early fibrous callus. ANIMALS AND METHODS: Calluses from rat rib fractures were removed 7, 14, and 21 days after fracture and unfractured ribs acted as controls. All tissues were analyzed using qPCR and immunohistochemistry. We analyzed expression of smooth muscle- and myofibroblast-associated genes and proteins including alpha smooth muscle actin (αSMA), non-muscle myosin, fibronectin extra domain A variant (EDA-fibronectin), OB-cadherin, connexin-43, basic calponin (h1CaP), and h-caldesmon. RESULTS: In calluses at 7 days post-fracture, there were statistically significant increases in expression of αSMA mRNA (2.5 fold), h1CaP mRNA (2.1 fold), EDA-fibronectin mRNA (14 fold), and connexin-43 mRNA (1.8 fold) compared to unfractured ribs, and by 21 days post-fracture mRNA expression in calluses had decreased to levels approaching those in unfractured rib. Immunohistochemistry of 7 day fibrous callus localized calponin, EDA-fibronectin and co-immunolabeling of OB-cadherin and αSMA (thus confirming a myofibroblastic phenotype) within various cell populations. INTERPRETATION: This study provides further evidence that early rat rib callus is not only smooth muscle-like in nature but also contains a notable population of cells that have a distinct myofibroblastic phenotype. The presence of these cells indicates that in vivo contraction of early callus is a mechanism that may occur in fractures so as to facilitate healing, as it does in soft tissue wound repair.