Lower Inflammatory Cell Infiltration Research Articles

Inflammatory response-based subtyping and potential therapeutic strategies for triple-negative breast cancer

Objective: Inflammatory response plays a crucial role in the development and treatment of cancer. However, the role of inflammatory response in triple-negative breast cancer (TNBC) remains unclear. Based on the heterogeneity of the inflammatory response, we classified TNBC, elucidated its subtype features, and revealed potential therapeutic strategies. Methods: We established inflammatory response subtyping based on the RNA sequencing data of TNBCs derived from a cohort at the Fudan University Shanghai Cancer Center (FUSCC). Next, we explored the features and potential therapeutic strategies for each subgroup by analyzing transcriptome data. Using a machine-learning method, we validated and generalized the TNBC inflammatory response subtypes in an external dataset. Results: A total of 360 TNBC samples and 88 normal tissues were collected from a cohort at FUSCC. Patients with TNBC were divided into four inflammatory response groups (IRGs) based on the expression of inflammatory response genes: high inflammatory response gene expression with pronounced pyroptosis phenotype and high immune cell infiltration (IRG 1), low inflammatory response gene expression and low immune cell infiltration (IRG 2), ITGB8 specific inflammatory response with a predominant proliferation phenotype (IRG 3), and low M1/M2 ratio with a marked angiogenesis phenotype (IRG 4). Relapse-free survival (RFS) was better in IRG 1 and 2 and worse in IRG 3 and 4. Owing to their poor prognosis, we mainly focused on IRG 3 and IRG 4 to investigate potential treatment strategies. ITGB8 was highly expressed in IRG 3; thus, targeting ITGB8 may be a potential therapeutic strategy for patients in IRG 3. IRG 4 had a lower M1/M2 ratio and a marked angiogenesis phenotype; therefore, therapeutic strategies, such as anti-angiogenesis or M2 to M1 repolarization of macrophages, could be recommended for these patients. Additionally, we validated and generalized the TNBC inflammatory response subtyping in an external dataset using a machine-learning method. Conclusion: TNBC patients with different inflammatory response subtypes have different characteristics and may need subtype-specific treatment strategies.

An Agonistic Monoclonal Antibody Targeting cMet Attenuates Inflammation and Up-Regulates Collagen Synthesis and Angiogenesis in Type 2 Diabetic Mice Wounds.

Diabetic wounds account for 25 to 50 percent of total diabetic health care costs annually, and present overall healing rates of less than 50 percent. Because delayed diabetic wound healing is associated with impaired fibroblast function, the authors hypothesize that tyrosine kinase Met (cMet) agonistic monoclonal antibody will promote diabetic wound healing by means of stable activation of hepatocyte growth factor/cMet signaling. Two 6-mm dorsal wounds were created in each mouse (6-week-old, male BKS.Cg-Dock7 m +/+Lepr db /J; n = 5). After subcutaneous injections of agonist (20 mg/kg) at 0 and 72 hours, the wound sizes were measured at days 0, 1, 3, 6, and 10. Histologic and immunohistochemical analyses were performed at day 10 (cMet, α-smooth muscle actin, CD68, and transforming growth factor-β). In vitro cytotoxicity and migration tests with diabetic fibroblasts were performed with or without agonist treatment (1 or 10 nM). cMet pathway activation of fibroblasts was confirmed through p-p44/42 mitogen-activated protein kinase, p-mTOR, p-cMet, and ROCK-1 expression. The cMet agonistic monoclonal antibody-treated group showed 1.60-fold lower wound area ( p = 0.027), 1.54-fold higher collagen synthesis ( p = 0.001), and 1.79-fold lower inflammatory cell infiltration ( p = 0.032) than the saline-treated control. The agonist increased cMet (1.86-fold; p = 0.029), α-smooth muscle actin (1.20-fold; p = 0.018), and vascular endothelial growth factor (1.68-fold, p = 0.029) expression but suppressed CD68 (1.25-fold; p = 0.043), transforming growth factor-β (1.25-fold; p = 0.022), and matrix metalloproteinase-2 (2.59-fold; p = 0.029) expression. In vitro agonist treatment (10 nM) of diabetic fibroblasts increased their migration by 8.98-fold ( p = 0.029) and activated the hepatocyte growth factor/cMet pathway. Tyrosine kinase Met agonistic monoclonal antibody treatment improved diabetic wound healing in mice and reduced wound-site inflammatory cell infiltration. These results need to be validated in large animals before piloting human trials. Although further clinical studies are necessary to evaluate its therapeutic efficacy, our study suggested that cMet agonistic monoclonal antibody can be the alternative modality in order to improve wound healing cascade in diabetic foot patients.

Early Intervention of Cold-Water Swimming on Functional Recovery and Spinal Pain Modulation Following Brachial Plexus Avulsion in Rats.

Brachial plexus avulsion (BPA) causes peripheral nerve injury complications with motor and sensory dysfunction of the upper limb. Growing evidence has shown an active role played by cold-water swimming (CWS) in alleviating peripheral neuropathic pain and functional recovery. This study examined whether CWS could promote functional recovery and pain modulation through the reduction of neuroinflammation and microglial overactivation in dorsal horn neurons at the early-stage of BPA. After BPA surgery was performed on rats, they were assigned to CWS or sham training for 5 min twice a day for two weeks. Functional behavioral responses were tested before and after BPA surgery, and each week during training. Results after the two-week training program showed significant improvements in BPA-induced motor and sensory loss (p < 0.05), lower inflammatory cell infiltration, and vacuole formation in injured nerves among the BPA–CWS group. Moreover, BPA significantly increased the expression of SP and IBA1 in dorsal horn neurons (p < 0.05), whereas CWS prevented their overexpression in the BPA–CWS group. The present findings evidenced beneficial rehabilitative effects of CWS on functional recovery and pain modulation at early-stage BPA. The beneficial effects are partially related to inflammatory suppression and spinal modulation. The synergistic role of CWS combined with other management approaches merits further investigation.

Berberine Influences the Survival of Fat Grafting by Inhibiting Autophagy and Apoptosis of Human Adipose Derived Mesenchymal Stem Cells.

ObjectiveHuman adipose-derived mesenchymal stem cells (ADSCs) have the potential to be applied to solid organ treatments. However, tissue regeneration is limited by the death of transplanted cells. Ischemia is the main cause of the poor outcome. This study aimed to investigate the effect of berberine (BBR) on ADSCs after fat grafting.MethodsThe antioxidant BBR on apoptosis and autophagy of ADSCs in vitro ischemia model was induced by hypoxia and serum deprivation (HY/SD). The autophagy promoter rapamycin and autophagy inhibitor 3-MA were incubated separately to investigate the crosstalk between autophagy and apoptosis. Pathway inhibitors further verified whether the autophagy and apoptosis were regulated by AMPK/mTor signaling pathway. Fat survival, fibrosis, level of inflammatory cell infiltration, and the effect of angiogenesis after BBR treatment were observed in vivo.ResultsBBR could reduce ROS production and reverse the decreasing cell survival rate. HY/SD would induce apoptosis and autophagy in ADSCs, and BBR could alleviate these processes. After interfering with the level of autophagy, we also proved that apoptosis was regulated by autophagy and changed accordingly. The results also indicated that BBR could protect against autophagy and apoptosis of ADSCs through AMPK/mTor pathway. The treated human-derived adipose tissue was transplanted into BALB/c nude mice, and with the intervention of BBR, the fat grafting had a higher survival rate, lower inflammatory cell infiltration and fibrosis level.ConclusionOur present study revealed that BBR was a promising anti-autophagy and apoptosis agent for improving the survival rate of ADSCs during cell transplantation.

Effects of protein extract from the housefly larvae (Musca domestica vicina) on the hyperlipidemic mice induced by Triton WR-1339

Hypercholesterolemia is a chief risk reason for atherosclerosis. Triton-WR 1339 (Tyloxapol) was used by several investigators to initiate hypercholesterolemia in animals. Housefly maggots contain high larval protein extract (PE) that is used as an anti-atherosclerosis. The goal of the current study was to evaluate the possible protective effects of the PE of houseflies against Triton-WR 1339-induced hyperlipidemia, oxidative damage, and histopathological changes of the liver and dorsal aorta of male mice. In this study, four groups of experimental mice, including ten mice each, were classified into: Group I, mice were administered with 0.5 ml saline solution, and considered as a positive control; Group II, mice were administered day after day by gavage with 0.5 ml of 200 mg/kg bw PE; Group III, hyperlipidemic mice were taken no treatment till the end of experiment and Group IV, the hyperlipidemic mice were administered day after day by gavage with 200 mg/kg bw PE. On day 40, the mice were anesthetized, blood samples were collected, the livers and the aorta were dissected for various biochemical tests and histopathological examinations. The biochemical parameters, triglycerides (TGs), total cholesterol (TC), low-density lipoprotein-cholesterol (LDL-C) and lipid peroxidation (LPO) were increased, and reduction in high-density lipoprotein-cholesterol (HDL-C), the total glutathione (GSH) and the glutathione peroxidase (GPx) levels were observed following Triton WR-1339 treatment in mice. After administration of the PE (200 mg/kg bw), the results revealed significant reduction in serum TC and TGs while HDL-C, oxidative stress levels in liver tissue, LPO, GSH and GPx were significantly increased. Histopathological examinations of the liver tissue and the aorta of mice showed alterations afterwards Triton WR-1339 administration. Treatment with 200 mg/kg bw PE showed beneficial effects with lower capsular thickening, enlargment of hepatocytes and much lower inflammatory cell infiltration.

Effects of rabbit pinna-derived blastema cells on tendon healing.

Objective(s):Tendon healing is substantially slow and often associated with suboptimal repair. Cell therapy is one of the promising methods to improve tendon repair. Blastema, a population of undifferentiated cells, represents characteristics of pluripotent mesenchymal stem cells and has the potentials to be used in regenerative medicine. The aim of this study was to investigate the use of blastema allotransplantation in rabbit tendon healing.Materials and Methods:In this study, one rabbit was used as a blastema donor, and twenty-four rabbits were divided into control and treatment groups. Blastema cells were obtained from ear pinna upon punch hole injury in the donor rabbit. Under general anesthesia, a complete transverse tenotomy was performed on the midsubstance of deep digital flexor tendon followed by suture-repair. In the treatment group, 1 × 106 blastema cells suspended in buffer saline were injected intratendinously at the repair site, while the control group received only the buffer saline. Cast coaptation was maintained for two weeks. Eight weeks after the operation, tendons were harvested, and histopathological, biomechanical, and biochemical assays were performed on samples. Results:Mechanical testing showed a significant increase in ultimate load, energy absorption, stiffness, yield load, stress, and strain in blastema-treated tendons compared to controls. Also, higher hydroxyproline content and improved collagen alignment along with lower inflammatory cell infiltration and decreased angiogenesis were observed in blastema-treated tendons. Conclusion:Increased levels of hydroxyproline and improved histopathological and biomechanical parameters in the treatment group suggest that blastema cells could be considered an adjunct to tendon repair in rabbits.

ECDI-fixed donor splenocytes prolong skin allograft survival by promoting M2 macrophage polarization and inducing regulatory T cells.

Rejection is a common complication of allogeneic tissue transplantation. Fixation of splenocytes (SP) with 1‐ethyl‐3‐(3'‐dimethylaminopropyl)‐carbodiimide (ECDI) induces immune tolerance in recipients post‐transplantation; however, the mechanism underlying this effect remains unclear. Here, we determined the mechanisms of ECDI‐fixed donor SP (ECDI‐SP) in inducing tolerance in skin allograft transplantation. C57BL/6‐recipient mice that received Balb/c full‐thickness skin transplants with two infusions of donor‐derived ECDI‐SP, along with rapamycin showed superior skin allograft survival and lower inflammatory cell infiltration than mice that received rapamycin‐only treatment. In ECDI‐SP‐treated mice, the levels of anti‐inflammatory cytokines such as interleukin (IL)‐10 in sera were markedly increased, whereas the expression of inflammatory cytokines was significantly suppressed. Splenic macrophages were significantly polarized to the alternative activated macrophage (M2) phenotype, with expansion of CD4+Foxp3+ regulatory T cells (Tregs) in the spleen and draining lymph nodes. Allostimulatory activity of ECDI‐SP in vitro and donor‐specific ex vivo hyporesponsiveness were observed. C57BL/6 macrophages engulfed allogeneic Balb/c‐derived ECDI‐SP, polarized to the M2 phenotype, with pronounced cAMP response element‐binding (CREB) protein phosphorylation. By facilitating increased IL‐10 expression, ECDI‐SP induced M2 polarization and Treg production, inhibiting effector T‐cell proliferation. Thus, ECDI‐SP modulates macrophage M2 polarization by increasing CREB phosphorylation and promoting Treg production to suppress allogeneic skin graft rejection.

Oral Phosphatidylcholine Improves Intestinal Barrier Function in Drug-Induced Liver Injury in Rats.

Objective Phosphatidylcholine (PC) is the major surface-active phospholipid and creates a hydrophobic nature to the surface. It has been reported to reverse the progression of liver fibrosis and to improve liver function. The aim of the present study was to evaluate the effects of orally administered PC on intestinal barrier function (IBF) in rats with drug-induced liver injury. Method Rats with carbon tetrachloride- (CCl4-) induced liver injury were treated with 100 mg/kg PC once daily for 21 days. The effects of PC therapy on (i) liver function and portal pressure, (ii) intestinal and hepatic histology, and (iii) plasma endotoxin, diamine oxidase (DAO), and tumour necrosis factor- (TNF-) α levels were investigated. Results PC therapy reduced portal pressure and improved the liver function in CCl4-induced liver injury. In PC-treated liver injury rats, collagen fibres were gradually decreased, while the disordered arrangement of hepatocytes and disorganized hepatic lobules were partially repaired, and inflammatory cell infiltration was decreased in the fibrous tissue. Lower inflammatory cell infiltration in the ileum improved intestinal histology, and reduced serum DAO levels were observed in PC-treated cirrhotic rats. These changes were associated with reduced inflammatory activity, as indicated by decreased serum TNF-α levels and plasma endotoxin levels. Conclusions These results suggest that PC therapy is hepatoprotective and is able to restore IBF and reduce endotoxaemia in rats with drug-induced liver injury.

Protective effects of alendronate in Triton X-100-induced hyperlipidemia in rats.

The aim of the present study was to evaluate the protective effects of alendronate (used in osteoporosis disease) in Triton X-100 (a polyethylene glycol-based non-ionic surfactant)-induced hyperlipidemia in rats. The animals were randomized into seven groups receiving different treatments for 21 days, and alendronate was administered (1.5 and 3 mg/kg body weight, per orally (p.o.) by oral gavage). On day 21, the rats were anesthetized and decapitated, blood samples were extracted, and the livers were dissected for various biochemical tests and histopathological examinations. The biochemical parameters, total cholesterol (TC), triglycerides (TGs), low-density lipoprotein-cholesterol (LDL-C), very low-density lipoprotein-cholesterol (VLDL-C), thiobarbituric acid reactive substances (TBARS), alanine aminotransferase (ALT), aspartate aminotransferase (AST), and atherogenic index, were increased, and reduction in high-density lipoprotein-cholesterol (HDL-C) levels was observed following Triton X-100 treatment to rats. Alendronate (1.5 and 3 mg/kg) produced a dose-dependent reduction in serum TC, VLDL-C, TGs, ratio of TC/HDL-C, ALT, AST, and TBARS. It significantly increased the HDL-C and superoxide dismutase levels but did not cause a significant decrease in serum LDL-C and/or an increase in catalase levels. Histopathological examinations of alendronate showed beneficial effects with lower capsular thickening, slight enlargement of the hepatocytes at the margin, and lower inflammatory cell infiltration. Alendronate showed dose-dependent antihyperlipidemic and hepatoprotective effects. It may serve a dual purpose as anti-osteoporotic and hypolipidemic by reducing blood cholesterol and TG synthesis and offering hepatic protection.

Novel Gut Barrier Modulatory Effect of Vildagliptin in Amelioration of Bacterial Translocation in Cholestatic Rats

Intestinal barrier dysfunction is frequently implicated in the etiology of septic complications following obstructive jaundice. The present study was conducted to investigate the potential gut barrier modulating effect of vildagliptin (VLD), a dipeptidyl peptidase‐IV (DPP‐IV) inhibitor, on common bile duct ligation (CBDL) in rats. VLD (10 mg/kg/d; p.o) was administered for 10 successive days after CBDL in male Sprague‐Dawley rats. VLD reduced portal endotoxemia induced by CBDL and suppressed rectal temperature elevation, body weight loss and colon and spleen indices to different extends. Moreover, it reversed villous blunting, low villous density, mucosal thinning and inflammatory cell infiltration noticed microscopically in the ileal segments of the CBDL rats. These events were accompanied by the reduction of intestinal myeloperoxidase activity and lipid peroxidation, the elevation of the major anti‐oxidant molecule, glutathione, as well as the normalization of ileal nitric oxide level. Furthermore, ileal DPP‐IV was inhibited in association with the rise in ileal glucagon‐like peptide‐1 (GLP‐1) and insulin‐like growth factor‐1 (IGF‐1) levels in VLD treated animals. The current findings suggest that VLD anti‐oxidant, anti‐inflammatory, as well as enhancements of incretin and IGF‐1 account for the hardwearing of the intestinal barrier that functions competently in the absence of the supporting bile elements.

Electronegative LDL is linked to high-fat, high-cholesterol diet–induced nonalcoholic steatohepatitis in hamsters

The pathogenesis of nonalcoholic steatohepatitis (NASH), like that of atherosclerosis, involves lipid accumulation, inflammation and fibrosis. Recent studies suggest that oxidized LDL (oxLDL) may be a risk factor for NASH, but oxLDL levels were not directly measured in these studies. The aim of this study was to examine whether there was an association between electronegative LDL [LDL(−)], a mildly oxLDL found in the blood, and the development of NASH using two animal models. Golden Syrian hamsters and C57BL/6 mice were fed a high-fat, high-cholesterol (HFC) diet for 6 or 12weeks, then liver lipid and histopathology, plasma lipoprotein profile and LDL(−) levels were examined. The HFC-diet-fed hamsters and mice had similar levels of hepatic lipid but different histopathological changes, with microvesicular steatosis, hepatocellular hypertrophy, inflammation and bridging fibrosis in the hamsters, but only in mild steatohepatitis with low inflammatory cell infiltration in the mice. It also resulted in a significant increase in plasma levels of LDL cholesterol and LDL(−) in hamsters, but only a slight increase in mice. Moreover, enlarged Kupffer cells, LDL(−) and accumulation of unesterified cholesterol were detected in the portal area of HFC-diet-fed hamsters, but not HFC-diet-fed mice. An in vitro study showed that LDL(−) from HFC-diet-fed hamsters induced TNF-α secretion in rat Kupffer cell through a LOX-1-dependent pathway. Our results strongly suggest that LDL(−) is one of the underlying causes of hepatic inflammation and plays a critical role in the development of NASH.

Treatment of Brain-Dead Donor Rats with Dimethyloxalylglycine Improves Both Donor and Graft Left-Ventricular Function after Heart Transplantation

Introduction: Cardiac transplantation is the most effective treatment of end-stage heart failure. Brain dead (BD) donors are currently the only reliable source for cardiac grafts. However, hemodynamic instability and cardiac dysfunction have been demonstrated in BD donors and this could therefore also affect posttransplant graft function. Hypothesis: Based upon the protective effect of Prolyl-Hydroxylase (PHD)-Hypoxia Inducible Factor (HIF)-1 pathway against ischemia/reperfusion injury (IRI), we tested the hypothesis that treatment of BD donor rats with the non-selective PHD inhibitor dimethyloxalylglycine (DMOG) results in a better graft heart condition in recipients after heterotopic transplantation. Methods: Lewis rats were injected with one shot DMOG (30 mg/kg i.v.) (n = 7) or equal volume of physiological saline (n = 7) and maintained BD for 5h by a subdurally placed and inflated balloon catheter. Controls were sham-operated (n = 11). Then, hearts were explanted, stored in cold preservation solution, heterotopically transplanted and after 1.5h reperfusion left-ventricular (LV) graft function was evaluated in vivo. Myocardial histological and molecular biological analyses were performed. Results: BD was associated with decreased LV function. DMOG treatment of BD animals resulted in better load-independent contractility and end-diastolic stiffness parameters (ESPVR E´max (mmHg/µl): BD+DMOG: 3.7 ± 0.6 vs BD: 3.1 ± 0.5; EDPVR (mmHg/µl): BD+DMOG: 0.13 ± 0.03 vs BD: 0.31 ± 0.06 p < 0.05). Following transplantation, DMOG treatment of BD donors significantly improved the altered LV systolic and diastolic function (at 80µL volume dP/dtmax (mmHg/s): BD+DMOG: 2284 ± 213 vs BD: 1854 ± 124 p < 0.05; dP/dtmin (mmHg/s): BD+DMOG: 1586 ± 183 vs BD: 1154 ± 74, p < 0.05; Tau-W (ms): BD+DMOG: 33 ± 4 vs BD: 43 ± 9 p < 0.05). Significantly lower myocardial inflammatory cell infiltration and necrosis were detected along with, significantly lower DNA-strand breakage in the DMOG treated BD group compared with BD group. Conclusions: Pre-treatment of BD heart donors with DMOG resulted in a significantly better LV graft function after transplantation. These results support the view that preconditioning of BD donors through the activation of HIF-1 pathway has a protective role against myocardial IRI after transplantation.

Using TP3DS as a surgical adjunct in oral and maxillofacial surgery

Background and objectives: Biocompatibility is a prerequisite for any substance to be used as an implant material, and potential biomaterials need to be characterized by histological evaluation of tissue responses. FullCure is an acrylic photopolymer with transparent appearance, manufactured by the Objet Company, used in 3-Dimensional Printing Systems for the manufacturing of stereolithography prototypes. Methods: The purpose of this study was to evaluate the bioactivity of FullCure compared to porous polyethylene implants (MedporR). To this end, tissue responses to both substances were evaluated in an animal model. Twelve FullCure laminae were applied on the right side of the dorsum of wistar rats (n= 6). In the same animals but in the left half was placed MedporeR. Responses and biocompatibility were assessed by histopathologic and histomorphometric evaluation at 3 months post-implantation. Results: FullCure showed marked biocompatibility. The implants were well integrated into the tissues: newly formed collagen bands were observed inside the implants and no granulomas were detected. FullCure implants showed a low inflammatory cell infiltration, without significative differences with respect to Medpor implants. Conclusions: FullCure is a biocompatible product that can be considered for use as a material implant in maxillofacial region. Further investigation is required to assess its long-term efficacy and safety.

OGG1, MYH and MTH1 gene variants identified in gastric cancer patients exhibiting both 8-hydroxy-2′-deoxyguanosine accumulation and low inflammatory cell infiltration in their gastric mucosa

Introduction 8-Hydroxy-2′-deoxyguanosine (8-OHdG) is one of the main DNA modifications produced by reactive oxygen species (ROS). Because 8-OHdG can pair with cytosine and adenine bases during DNA synthesis, when 8-hydroxyguanine (8-OHG) is present in the DNA template, it causes G:C to T:A transversions (Shibutani et al. 1991), and it induces A:T to C:G transversions when 8-hydroxy-dGTP in the nucleotide pool is incorporated into DNA (Maki and Sekiguchi 1992). Because of these transversions, 8-OHdG accumulation is thought to cause carcinogenesis. 8-OHdG accumulation in mammalian cells is prevented by the base excision repair enzymes OGG1, MYH and NEIL1, and by MTH1, an enzyme that removes 8-hydroxy-dGTP from the intracellular nucleotide pool (Nakabeppu 2001). A variety of factors, including sodium chloride, Helicobacter pylori infection and smoking (Tredaniel et al. 1997; Farinati et al. 1998), induce inflammation in the stomach tissue. A considerable inflammatory cell infiltrate in the gastric mucosa causes the production of ROS (Ernst 1999), and ROS are thought to lead to 8-OHdG accumulation in the gastric mucosa (Farinati et al. 1998), suggesting that the level of inflammatory cell infiltration in the stomach may be one of the factors that determine the 8-OHdG level in the stomach. We, therefore, hypothesized that gastric cancer patients with both 8-OHdG accumulation and low level of inflammatory cell infiltration in their stomach have genetic factors that cause them to have a low ability to repair 8-OHdG. We selected 23 patients exhibiting mild or no neutrophil and mono-

Neurogenic Factors in the Impaired Healing of Diabetic Foot Ulcers

We hypothesize that the reduced innervation of skin can be observed both in clinically neuropathic and non-neuropathic diabetic foot ulcers and can contribute to low inflammatory cell infiltration. Twenty patients with type 2 diabetes and active foot ulcers, without clinical evidence of peripheral sensory neuropathy (n = 12) and with sensory neuropathy (n = 8) were involved in this study. Biopsies from ulcer margin were examined immunohistochemically. Studies revealed presence of protein gene product 9.5 (PGP9.5)+ nerve endings only in reticular dermis in 3 of 12 non-neuropathic subjects, however, regenerating GAP-43+ endings were seen in dermis of almost all specimens. Lack of substance P+ nerve endings was characteristic for both groups. The reduced distribution of calcitonin gene-related peptide+ nerves in epidermis and dermis was seen mainly in neuropathic group. In neo-epidermis lack of nerve growth factor expression was observed in both groups, whereas neurotrophin 3 immunostaining was characteristic for neuropathic specimens (P < 0.03). Expression of trkA and trkC receptors did not differ significantly between groups. Low inflammatory cell infiltration and moderate presence of fibroblasts was characteristic for all studied specimens. The observed reduction of foot skin innervation and neurogenic factors expression can be correlated with low inflammatory cell accumulation and subsequently leads to the observed chronicity of diabetic foot ulcer healing process in both neuropathic and non-neuropathic patients.

Amelioration of Collagen-Induced Arthritis in Mice by a Saponin Fraction from Gleditsia sinensis.

In the current study, we investigated the therapeutic potential and underlying mechanisms of the saponin fraction from anomalous fruits of Gleditsia sinensis. Lam. (Leguminosae) (SFGS) on collagen II (CII)-induced arthritis (CIA) in DBA/1 mice. SFGS (50, 100, and 200 mg/kg), orally administered from the day of immunization, dose-dependently alleviated disease severity, postponed the onset and reduced the incidence rate of CIA. Histologic analysis revealed that joints of CIA mice treated with SFGS showed low inflammatory cell infiltration and slight synovium hyperplasia and focal bone erosion. Furthermore, SFGS treatments lowered the serum anti-CII autoantibody levels and suppressed the delayed-type hypersensitivity against CII in the ears of CIA mice. The findings indicated that SFGS ameliorated inflammation and joint destruction in CIA mice, which may be the consequence of suppression on CII-specific humoral and cellular immunity. SFGS should be a candidate novel therapeutic agent for rheumatoid arthritis.

Adenovirus-Mediated Gene Transfer of Superoxide Dismutase and Catalase Decreases Restenosis after Balloon Angioplasty

Background: Reactive oxygen species (ROS) production increases after injury and potentially contributes to restenosis after angioplasty. We therefore evaluated the effect of adenovirus-mediated gene transfer (Ad) of superoxide dismutase (SOD) and catalase (CAT) on ROS production and restenosis after balloon angioplasty. Methods: O₂^– and H₂O₂production was quantified in cultured cells after incubation with either LPS or CuSO₄. Angioplasty and gene transfer were performed in rabbit atherosclerotic iliac arteries. One artery was injected with AdSOD and AdCAT, while the contralateral artery was injected with an adenovirus carrying no transgene, and served as control. Results: ROS production was significantly decreased after adenovirus-mediated gene transfer of SOD and CAT as compared with control. Treated arteries showed less restenosis (32 ± 27 vs. 63 ± 19%, p = 0.003) and less constrictive remodeling (1.2 ± 0.3 vs. 0.9 ± 0.2, p = 0.02) than control arteries. Arteries injected with AdSOD and AdCAT showed better vasoreactivity to acetylcholine (11 ± 4 vs. –1 ± 6%, p < 0.05), lower collagen density (43 ± 16 vs. 53 ± 23%, p = 0.03), and lower inflammatory cell infiltration (22 ± 6 vs. 36 ± 11%, p = 0.04) than control arteries. Conclusions: Our data suggest that adenovirus-mediated gene transfer of SOD and CAT reduced oxidative stress, restenosis, collagen accumulation, and inflammation and improved endothelial function after angioplasty.

Increased gene expression and inflammatory cell infiltration caused by electroporation are both important for improving the efficacy of DNA vaccines

One potential reason for the enhancement of immune responses to DNA vaccines following electroporation is increased gene expression. However, the inflammatory response and accompanying cellular infiltration stimulated by electroporation may also be essential for enhancing immune responses to DNA vaccines. These parameters were investigated in pigs, using different electroporation conditions to induce different levels of gene expression and inflammation. Results indicated that the least effective strategy was conventional intramuscular injection where there was low gene expression and low inflammatory cell infiltration. The most efficacious strategy was plasmid administration immediately followed by electroporation. This latter set of conditions elicited a combination of high gene expression and high cellular infiltration. This indicates that electroporation enhances immune responses to DNA vaccines through increased gene expression and inflammatory cell infiltration.