Functional CRISPR dissection of gene networks controlling human regulatory T cell identity.

Human regulatory T (Treg) cells are essential for immune homeostasis. The transcription factor FOXP3 maintains Treg cell identity, yet the complete set of key transcription factors that control Treg cell gene expression remains unknown. Here, we used pooled and arrayed Cas9 ribonucleoprotein screens to identify transcription factors that regulate critical proteins in primary human Treg cells under basal and proinflammatory conditions. We then generated 54,424 single-cell transcriptomes from Treg cells subjected to genetic perturbations and cytokine stimulation, which revealed distinct gene networks individually regulated by FOXP3 and PRDM1, in addition to a network coregulated by FOXO1 and IRF4. We also discovered that HIVEP2, to our knowledge not previously implicated in Treg cell function, coregulates another gene network with SATB1 and is important for Treg cell-mediated immunosuppression. By integrating CRISPR screens and single-cell RNA-sequencing profiling, we have uncovered transcriptional regulators and downstream gene networks in human Treg cells that could be targeted for immunotherapies.

CRISPR screens decode cancer cell pathways that trigger γδ T cell detection.

γδ T cells are potent anticancer effectors with the potential to target tumours broadly, independent of patient-specific neoantigens or human leukocyte antigen background1-5. γδ T cells can sense conserved cell stress signals prevalent in transformed cells2,3, although the mechanisms behind the targeting of stressed target cells remain poorly characterized. Vγ9Vδ2 T cells-the most abundant subset of human γδ T cells4-recognize a protein complex containing butyrophilin 2A1 (BTN2A1) and BTN3A1 (refs. 6-8), a widely expressed cell surface protein that is activated by phosphoantigens abundantly produced by tumour cells. Here we combined genome-wide CRISPR screens in target cancer cells to identify pathways that regulate γδ T cell killing and BTN3A cell surface expression. The screens showed previously unappreciated multilayered regulation of BTN3A abundance on the cell surface and triggering of γδ T cells through transcription, post-translational modifications and membrane trafficking. In addition, diverse genetic perturbations and inhibitors disrupting metabolic pathways in the cancer cells, particularly ATP-producing processes, were found to alter BTN3A levels. This induction of both BTN3A and BTN2A1 during metabolic crises is dependent on AMP-activated protein kinase (AMPK). Finally, small-molecule activation of AMPK in a cell line model and in patient-derived tumour organoids led to increased expression of the BTN2A1-BTN3A complex and increased Vγ9Vδ2 T cell receptor-mediated killing. This AMPK-dependent mechanism of metabolic stress-induced ligand upregulation deepens our understanding of γδ T cell stress surveillance and suggests new avenues available to enhance γδ T cell anticancer activity.

Obesity alters pathology and treatment response in inflammatory disease.

Decades of work have elucidated cytokine signalling and transcriptional pathways that control T cell differentiation and have led the way to targeted biologic therapies that are effective in a range of autoimmune, allergic and inflammatory diseases. Recent evidence indicates that obesity and metabolic disease can also influence the immune system1-7, although the mechanisms and effects on immunotherapy outcomes remain largely unknown. Here, using two models of atopic dermatitis, we show that lean and obese mice mount markedly different immune responses. Obesity converted the classical type 2 T helper (TH2)-predominant disease associated with atopic dermatitis to a more severe disease with prominent TH17 inflammation. We also observed divergent responses to biologic therapies targeting TH2 cytokines, which robustly protected lean mice but exacerbated disease in obese mice. Single-cell RNA sequencing coupled with genome-wide binding analyses revealed decreased activity of nuclear receptor peroxisome proliferator-activated receptor-γ (PPARγ) in TH2 cells from obese mice relative to lean mice. Conditional ablation of PPARγ in T cells revealed that PPARγ is required to focus the in vivo TH response towards a TH2-predominant state and prevent aberrant non-TH2 inflammation. Treatment of obese mice with a small-molecule PPARγ agonist limited development of TH17 pathology and unlocked therapeutic responsiveness to targeted anti-TH2 biologic therapies. These studies reveal the effects of obesity on immunological disease and suggest a precision medicine approach to target the immune dysregulation caused by obesity.

The effect of age on alpha rhythms in the human brain derived from source localized resting-state electroencephalography.

With increasing age, peak alpha frequency (PAF) is slowed, and alpha power is reduced during resting-states with eyes closed. These age-related changes are evident across the whole scalp but remained unclear at the source level. The purpose of this study was to determine whether age impacts the power and frequency of the dominant alpha rhythm equally across source generators or whether the impact of age varies across sources. A total of 28 young adults and 26 elderly adults were recruited. High-density EEG was recorded for 10 mins with eyes closed. Single dipoles for each independent component were localized and clustered based on their anatomical label, resulting in 36 clusters. Meta-analyses were then conducted to assess effect sizes for PAF and power at PAF for all 36 clusters. Subgroup analyses were then implemented for frontal, sensorimotor, parietal, temporal, and occipital regions. The results of the meta-analyses showed that the elderly group exhibited slower PAF and less power at PAF compared to the young group. Subgroup analyses revealed age effects on PAF in parietal (g = 0.38), temporal (g = 0.65), and occipital regions (g = 1.04), with the largest effects observed in occipital regions. For power at PAF, age effects were observed in sensorimotor (g = 0.84) and parietal regions (g = 0.80), with the sensorimotor region showing the largest effect. Our findings show that age-related slowing and attenuation of the alpha rhythm manifests differentially across cortical regions, with sensorimotor and occipital regions most susceptible to age effects.

RGC-32 mediates proinflammatory and profibrotic pathways in immune-mediated kidney disease.

Systemic lupus erythematosus is an autoimmune disease that results in immune-mediated damage to kidneys and other organs. We investigated the role of response gene to complement-32 (RGC-32), a proinflammatory and profibrotic mediator induced by TGFß and C5b-9, in nephrotoxic nephritis (NTN), an experimental model that mimics human lupus nephritis. Proteinuria, loss of renal function and kidney histopathology were attenuated in RGC-32 KO NTN mice. RGC-32 KO NTN mice displayed downregulation of the CCL20/CCR6 and CXCL9/CXCR3 ligand/receptor pairs resulting in decreased renal recruitment of IL-17+ and IFNγ+ cells and subsequent decrease in the influx of innate immune cells. RGC-32 deficiency attenuated renal fibrosis as demonstrated by decreased deposition of collagen I, III and fibronectin. Thus, RGC-32 is a unique mediator shared by the Th17 and Th1 dependent proinflammatory and profibrotic pathways and a potential novel therapeutic target in the treatment of immune complex mediated glomerulonephritis such as lupus nephritis.

Interfacing High-Throughput Electrosynthesis and Mass Spectrometric Analysis of Azines.

Combinatorial electrochemistry has great promise for accelerated reaction screening, organic synthesis, and catalysis. Recently, we described a new high-throughput electrochemistry platform, colloquially named "Legion". Legion fits the footprint of a 96-well microtiter plate with simultaneous individual control over all 96 electrochemical cells. Here, we demonstrate the versatility of Legion when coupled with high-throughput mass spectrometry (MS) for electrosynthetic product screening and quantitation. Electrosynthesis of benzophenone azine was selected as a model reaction and was arrayed and optimized using a combination of Legion and nanoelectrospray ionization MS. The combination of high-throughput synthesis with Legion and analysis via MS proves a compelling strategy for accelerating reaction discovery and optimization in electro-organic synthesis.

The landscape of liver transplantation for patients with alcohol-associated liver disease in the United States.

Indications for liver transplants have expanded to include patients with alcohol-associated liver disease (ALD) over the last decade. Concurrently, the liver allocation policy was updated in February 2020 replacing the Donor Service Area with Acuity Circles (ACs). The aim is to compare the transplantation rate, waitlist outcomes, and posttransplant survival of candidates with ALD to non-ALD and assess differences in that effect after the implementation of the AC policy. Scientific Registry for Transplant Recipients data for adult candidates for liver transplant were reviewed from the post-AC era (February 4, 2020-March 1, 2022) and compared with an equivalent length of time before ACs were implemented. The adjusted transplant rates were significantly higher for those with ALD before AC, and this difference increased after AC implementation (transplant rate ratio comparing ALD to non-ALD = 1.20, 1.13, 1.61, and 1.32 for the Model for End-Stage Liver Disease categories 37-40, 33-36, 29-32, and 25-28, respectively, in the post-AC era, p < 0.05 for all). The adjusted likelihood of death/removal from the waitlist was lower for patients with ALD across all lower Model for End-Stage Liver Disease categories (adjusted subdistribution hazard ratio = 0.70, 0.81, 0.84, and 0.70 for the Model for End-Stage Liver Disease categories 25-28, 20-24, 15-19, 6-14, respectively, p < 0.05). Adjusted posttransplant survival was better for those with ALD (adjusted hazard ratio = 0.81, p < 0.05). Waiting list and posttransplant mortality tended to improve more for those with ALD since the implementation of AC but not significantly. ALD is a growing indication for liver transplantation. Although patients with ALD continue to have excellent posttransplant outcomes and lower waitlist mortality, candidates with ALD have higher adjusted transplant rates, and these differences have increased after AC implementation.

Expression and characterization of a lipase EstA from Bacillus subtilis KM-BS for application in bio-hydrolysis of waste cooking oil.

A lipase EstA from Bacillus subtilis KM-BS was expressed in Escherichia coli BL21 (DE3) cells. The recombinant enzyme achieved high activity (49.67 U/mL) with protein concentration of 1.29 mg/mL under optimal conditions at the large-scale expression of 6 h and post-induction time at 30 °C using 0.1 mM isopropyl-ß-d-thiogalactopyranoside (IPTG). The optimal temperature and pH of the purified enzyme were at 45-55 °C and pH 8.0 - 9.0, respectively. Activity of the purified enzyme was stable in the presence of 1 mM Ca2+; stimulated by 1 mM Mg2+ and Mn2+, and inhibited by Fe3+. A significant amount of fatty acids was released during the hydrolysis of waste cooking oil under the catalysis of purified lipase, indicating that this recombinant lipase showed promise as a suitable candidate in industrial fields, particularly in biodiesel and detergent sector.

Synthesis of a 3D Flower-Like BiOCl/Bi-MOF Heterostructure for High-Performance Removal of Rhodamine B and Tetracycline Hydrochloride.

Semiconductor materials based on bismuth metal have been extensively explored for their potential in photocatalytic applications owing to their distinctive crystal structure. Herein, we present the development of a hybrid photocatalyst, CAU-17/BiOCl, featuring a flower-like nanosheet morphology tailored for the photocatalytic degradation of organic contaminants such as rhodamine B (RhB) and tetracycline hydrochloride (TCH). The composite material is obtained by growing thin CAU-17 layers directly onto the host flower-like BiOCl nanosheets under solvothermal conditions. The optimized CAU-17/BiOCl composite possesses excellent photocatalytic performance, achieving a notable 96.0% removal rate for RhB and 78.4% for TCH after 60 and 90 min of LED light irradiation, respectively. This boosted activity is attributed to the heightened absorption of visible light caused by BiOCl and the provision of additional reaction sites due to the thin CAU-17 layers. Furthermore, the establishment of an S-scheme heterojunction mechanism enables efficient charge separation between CAU-17 and BiOCl, facilitating the separation of photoinduced electrons (e-) and holes (h+). Analysis of the degradation mechanism of RhB and TCH reveals the predominant role of superoxide radicals (•O2-), e-, and h+ in the photocatalytic degradation process. Moreover, the removal efficiency of TCH can reach approximately 64.5% after four cycles of recycling of CAU-17/BiOCl. Our work provides a facile, effective solution and a theoretically explained approach for the effective degradation of pollutants using heterojunction photocatalysts.

Anterior approach for a stab wound with penetrating rebar injury causing incomplete cauda equina syndrome in lumbosacral spine lesion: a case report and literature review.

PURPOSE: To report a unique case of incomplete CES following a rebar penetrating injury in perineal region with retro-pulsed fragment, which was treated with anterior approach and discuss suitable surgical approach. METHODS: Incomplete cauda equina syndrome caused by non-missile penetrating injury is extremely rare. A 26-year-old male patient presented incomplete cauda equina syndrome due to a penetrating rebar wound from his perineal region to the lumbosacral spine. Computed tomography demonstrated a bony fragment broken from S1 body compressing into the spinal canal. RESULTS: By anterior approach, we performed partial corpectomy of L5, decompression by retrieving the bony fragment and L5-S1 interbody fusion. The patient had a significant recovery, and no clinical complication was found after over 2-year follow-up. CONCLUSION: It is challenging to determine the optimal strategy of surgical treatment for penetrating spinal injuries with retained foreign bodies, here we suggest an anterior approach situation that has the advantage of being able to effectively perform decompression and prevent iatrogenic damages of thecal sac and nerve rootlets.

A transformer architecture for retention time prediction in liquid chromatography mass spectrometry-based proteomics.

Accurate retention time (RT) prediction is important for spectral library-based analysis in data-independent acquisition mass spectrometry-based proteomics. The deep learning approach has demonstrated superior performance over traditional machine learning methods for this purpose. The transformer architecture is a recent development in deep learning that delivers state-of-the-art performance in many fields such as natural language processing, computer vision, and biology. We assess the performance of the transformer architecture for RT prediction using datasets from five deep learning models Prosit, DeepDIA, AutoRT, DeepPhospho, and AlphaPeptDeep. The experimental results on holdout datasets and independent datasets exhibit state-of-the-art performance of the transformer architecture. The software and evaluation datasets are publicly available for future development in the field.

Neu(t)ralMC: energy-efficient open source Monte Carlo algorithm for assessing photon transport in turbid media.

Light propagation in turbid mediums such as atmosphere, fluids, and biological tissues is a challenging problem which necessitates accurate simulation techniques to account for the effects of multiple scattering. The Monte Carlo method has long established itself as a gold standard and is widely adopted for simulating light transport, however, its computationally intensive nature often requires significant processing power and energy consumption. In this paper a novel, open source Monte Carlo algorithm is introduced which is specifically designed for use with energy-efficient processors, effectively addressing those challenges, while maintaining the accuracy/compatibility and outperforming existing solutions. The proposed implementation optimizes photon transport simulations by exploiting the unique capabilities of Apple's low-power, high-performance M-family of chips. The developed method has been implemented in an open-source software package, enabling seamless adaptation of developed algorithms for specific applications. The accuracy and performance are validated using comprehensive comparison with existing solvers commonly used for biomedical imaging. The results demonstrate that the new algorithm achieves comparable accuracy levels to those of existing techniques while significantly reducing computational time and energy consumption.

The SWI/SNF Complex: A Frequently Mutated Chromatin Remodeling Complex in Cancer.

The switch/sucrose non-fermenting (SWI/SNF) chromatin remodeling complex is a global regulator of gene expression known to maintain nucleosome-depleted regions at active enhancers and promoters. The mammalian SWI/SNF protein subunits are encoded by 29 genes and 11-15 subunits including an ATPase domain of either SMARCA4 (BRG1) or SMARCA2 (BRM) are assembled into a complex. Based on the distinct subunits, SWI/SNF are grouped into 3 major types (subfamilies): the canonical BRG1/BRM-associated factor (BAF/cBAF), polybromo-associated BAF (PBAF), and non-canonical BAF (GBAF/ncBAF). Pan-cancer genome sequencing studies have shown that nearly 25% of all cancers bear mutations in subunits of the SWI/SNF complex, many of which are loss of function (LOF) mutations, suggesting a tumor suppressor role. Inactivation of SWI/SNF complex subunits causes widespread epigenetic dysfunction, including increased dependence on antagonistic components such as polycomb repressor complexes (PRC1/2) and altered enhancer regulation, likely promoting an oncogenic state leading to cancer. Despite the prevalence of mutations, most SWI/SNF-mutant cancers lack targeted therapeutic strategies. Defining the dependencies created by LOF mutations in SWI/SNF subunits will identify better targets for these cancers.

FAS and SREBP-1c Inhibition via AMPK Activation in HepG2 Cells by Biovip, Tox-off, and Traphanoside GO1 from Glinus oppositifolius.

Glinus oppositifolius is an endemic herbaceous plant found in tropical Asian countries and is native in Vietnam. It is used in traditional folk medicine because of its flavor and antiseptic and laxative effects. In the current research, the effects of Tox-off, Biovip, and the purified compounds isolated from G. oppositifolius in the previous study were evaluated on the activation of adenosine 5'-monophosphate-activated protein kinase (AMPK)-activated protein kinase (AMPK) and acetyl-coenzyme A carboxylase (ACC) in C2C12 myoblasts. In addition, the most potent active compounds, traphanoside-GO1 (TRA-GO1) and TRA-GO5 have validated the reduction of fatty acid synthase (FAS) and sterol regulatory element binding protein (SREBP)-1c in HepG2 cells. We found that Tox-off and Biovip significantly increased the phosphorylation of AMPK and ACC in C2C12 myoblasts. Furthermore, TRA-GO1 and TRA-GO5 significantly increased the AMPK activation and phosphorylation of its downstream substrate ACC in a concentration-dependent way compared to the dimethyl sulfoxide (DMSO) control. Besides, the protein level of FAS and SREBP-1c decreased by TRA-GO1 and TRA-GO5 in a concentration-dependent manner. Taken together, our results showed that the increased AMPK and ACC phosphorylation by active components of G. oppositifolius may activate the AMPK signaling pathways, which are useful for the anti-obesity and its related metabolic disorders.

Analysing Monday discharges to identify lost opportunities for weekend discharge.

Lower rates of hospital discharge occur on weekends compared with weekdays. The authors performed a retrospective chart review of Monday discharges from the Hospital Medicine service at an academic hospital over a 3-month period to identify reasons for delayed discharge despite medical stability. Of 202 eligible patients, 81 (40%) had documentation indicating stability for earlier discharge. Common causes included bed availability or insurance authorisation at a skilled nursing facility, home care services and patient/family disagreement with discharge.

Dapagliflozin for heart failure according to body mass index: the DELIVER trial.

AIMS: Obesity is common and associated with unique phenotypic features in heart failure with preserved ejection fraction (HFpEF). Therefore, understanding the efficacy and safety of new therapies in HFpEF patients with obesity is important. The effects of dapagliflozin were examined according to body mass index (BMI) among patients in the Dapagliflozin Evaluation to Improve the LIVEs of Patients With PReserved Ejection Fraction Heart Failure trial. METHODS AND RESULTS: Body mass index was analysed by World Health Organization (WHO) categories and as a continuous variable using restricted cubic splines. Body mass index ranged from 15.2 to 50 kg/m2 with a mean value of 29.8 (standard deviation ± 6.1) kg/m2. The proportions, by WHO category, were: normal weight 1343 (21.5%); overweight 2073 (33.1%); Class I obesity 1574 (25.2%); Class II obesity 798 (12.8%); and Class III obesity 415 (6.6%). Compared with placebo, dapagliflozin reduced the risk of the primary outcome to a similar extent across these categories: hazard ratio (95% confidence interval): 0.89 (0.69-1.15), 0.87 (0.70-1.08), 0.74 (0.58-0.93), 0.78 (0.57-1.08), and 0.72 (0.47-1.08), respectively (P-interaction = 0.82). The placebo-corrected change in Kansas City Cardiomyopathy Questionnaire total symptom score with dapagliflozin at 8 months was: 0.9 (-1.1, 2.8), 2.5 (0.8, 4.1), 1.9 (-0.1, 3.8), 2.7 (-0.5, 5.8), and 8.6 (4.0, 13.2) points, respectively (P-interaction = 0.03). The placebo-corrected change in weight at 12 months was: -0.88 (-1.28, -0.47), -0.65 (-1.04, -0.26), -1.42 (-1.89, -0.94), -1.17 (-1.94, -0.40), and -2.50 (-4.4, -0.64) kg (P-interaction = 0.002). CONCLUSIONS: Obesity is common in patients with HFpEF and is associated with higher rates of heart failure hospitalization and worse health status. Treatment with dapagliflozin improves cardiovascular outcomes across the spectrum of BMI, leads to greater symptom improvement in patients with obesity, compared with those without, and has the additional benefit of causing modest weight loss.

Response Methodology Optimization and Artificial Neural Network Modeling for the Removal of Sulfamethoxazole Using an Ozone-Electrocoagulation Hybrid Process.

Removing antibiotics from water is critical to prevent the emergence and spread of antibiotic resistance, protect ecosystems, and maintain the effectiveness of these vital medications. The combination of ozone and electrocoagulation in wastewater treatment provides enhanced removal of contaminants, improved disinfection efficiency, and increased overall treatment effectiveness. In this work, the removal of sulfamethoxazole (SMX) from an aqueous solution using an ozone-electrocoagulation (O-EC) system was optimized and modeled. The experiments were designed according to the central composite design. The parameters, including current density, reaction time, pH, and ozone dose affecting the SMX removal efficiency of the OEC system, were optimized using a response surface methodology. The results show that the removal process was accurately predicted by the quadric model. The numerical optimization results show that the optimum conditions were a current density of 33.2 A/m2, a time of 37.8 min, pH of 8.4, and an ozone dose of 0.7 g/h. Under these conditions, the removal efficiency reached 99.65%. A three-layer artificial neural network (ANN) with logsig-purelin transfer functions was used to model the removal process. The data predicted by the ANN model matched well to the experimental data. The calculation of the relative importance showed that pH was the most influential factor, followed by current density, ozone dose, and time. The kinetics of the SMX removal process followed the first-order kinetic model with a rate constant of 0.12 (min-1). The removal mechanism involves various processes such as oxidation and reduction on the surface of electrodes, the reaction between ozone and ferrous ions, degradation of SMX molecules, formation of flocs, and adsorption of species on the flocs. The results obtained in this work indicate that the O-EC system is a potential approach for the removal of antibiotics from water.

Comparison of Implant Stability Between Conventional Drilling and Piezosurgical Implant Bed Preparation Techniques.

This prospective study compared the stability of implants placed using piezoelectric surgery (piezo group) and those placed using conventional rotary drills (bur group) during the first 90 days postoperatively. Teeth in the posterior maxillary regions of 21 patients were randomly assigned to 2 groups. The implant stability quotient (ISQ) was measured at days 0, 7, 14, 21, 28, 42, 56, and 90 postoperatively. Twenty-eight of 29 implants were successfully integrated at day 90 (1 implant in the test group was lost). Although both groups showed a significant overall increase in implant stability with time (P < .0001) and a high final mean ISQ value, no statistically significant difference in stability was seen between the groups. The bur group showed greater variance in ISQ values than the piezo group did (P < .001) at all time points. Long-term studies with larger samples are needed to investigate the bone response to the use of piezoelectric surgery for implant preparation.

Drug-Related Problems and Pharmacists' Interventions in Hypertensive Outpatients: A Multicenter Prospective Study in 3 Vietnamese Hospitals.

Background: Clinical pharmacists' interventions (PIs) on drug-related problems (DRPs) in Vietnamese hypertensive outpatients are limited. Objectives: The objective was to investigate the prevalence and nature of DRPs, and factors which are likely to have DRPs, types of PIs, and their acceptance rate in 3 Vietnamese hospitals. Method: A prospective interventional study was conducted over a period of 3 months in 3 hospitals (from October 2021 to March 2022). Clinical pharmacists conducted medication reviews after collecting patient information from prescriptions and patient interviewing, and then identified the DRPs and suggested PIs according to the Vi-Med tool. These DRPs and PIs were reviewed by other superior clinical pharmacists and a consensus meeting with 3 cardiologists. Results: Of 381 patients included, 344 (90.23%) experienced 1 or more DRPs. A total of 820 DRPs were identified with an average of 2.15 DRPs per patient and 415 (50.61%) were hypertension-related issues. The most common DRPs identified were "administration mode" (46.34%), "missing indication" (18.05%), "non-conformity indication" (17.80%), and "dosage" (11.95%). Comorbidity (adjusted odds ratio [AOR] = 3.985, 95% CI: 1.597-9.942, P = 0.003) was the predictor of DRPs. Clinical pharmacists provided 739 PIs and 94.45% were accepted by physicians. Conclusion: The results of this study showed that DRPs were very common in hypertensive outpatients and highlighted the role of clinical pharmacists to identify and resolve DRPs through prompt interventions.

RGC-32' dual role in smooth muscle cells and atherogenesis.

Proliferation of endothelial cells (EC) and smooth muscle cells (SMC) is a critical process in atherosclerosis. Here, we investigated the involvement of sublytic C5b-9 effector Response Gene to Complement 32 (RGC-32) in cell cycle activation, phenotypic switch, and production of extracellular matrix (ECM) in SMC. Overexpression of RGC-32 augmented C5b-9-induced cell cycle activation and proliferation of SMC in an ERK1-dependent manner and silencing of RGC-32 inhibited C5b-9-induced cell cycle activation. C5b-9-induced cell cycle activation also required phosphorylation of RGC-32 at threonine 91. We found that ECM components fibronectin and collagens I-V were expressed by SMC in human aortic atherosclerotic tissue. Silencing of RGC-32 in cultured SMC was followed by a significant reduction in TGF-ß-induced expression of SMC differentiation markers myocardin, SM22 and α-SMA, and that of collagens I, IV and V. These data suggest that RGC-32 participates in both sublytic C5b-9-induced cell cycle activation and TGF-ß-induced ECM production.