Mannan-binding lectin reduces CpG DNA-induced inflammatory cytokine production by human monocytes.

Mannan-binding lectin (MBL) belongs to the collectin family and functions as an opsonin that can also initiate complement activation. Our previous study showed that MBL serves as a double-stranded RNA binding protein that attenuates polyriboinosinic-polyribocytidylic acid-induced TLR3 activation. Prompted by these findings, in the present study cross-talk between MBL and CpG-DNA-induced TLR9 activation was investigated. Here, it was found that MBL also interacts with the TLR9 agonist, CpG oligodeoxynucleotide (CpG-ODN), in a calcium-dependent manner. Purified MBL protein suppressed activation of nuclear factor-kappa B signaling and subsequent production of proinflammatory cytokines from human monocytes induced by CpG-ODN 2006. These observations indicate that MBL can down-regulate CpG DNA-induced TLR9 activation, emphasizing the importance of understanding the interaction of MBL with TLR agonist in host immune defense.

Mannan-binding lectin at supraphysiological concentrations inhibits differentiation of dendritic cells from human CD14+ monocytes.

Mannan-binding lectin (MBL), a circulating C-type lectin, is an important member of the defense collagen family. It exhibits a high potential for recognizing broad categories of pathogen-associated molecular patterns and initiating complement cascade responses. DCs are well-known specialist antigen-presenting cells that significantly trigger specific T cell-mediated immune responses. In our previous study, it was observed that high concentrations of MBL significantly attenuate LPS-induced maturation of monocyte-derived DCs (MoDCs). In the current study, it was postulated that MBL at similar supraphysiological concentrations would affect early differentiation of MoDCs in some way. CD14(+) monocytes from human peripheral blood mononuclear cells were cultured with granulocyte-macrophage colony-stimulating factor and IL-4 in the presence or absence of physiological (1 µg/mL) and supraphysiological concentrations (20 µg/mL) of MBL protein, respectively. Phenotypic analysis indicated that the differentiated DCs incubated with high concentrations of MBL expressed MHC class II and costimulatory molecules (e.g., CD80 and CD40) more weakly than did control groups. The secretion of IL-10 and IL-6 increased markedly, whereas their mixed lymphocyte reaction-stimulating capacity decreased. Members of the signal transducer and activator of transcription family were also found to be differentially regulated. Thus, beyond the role of MBL as an opsonin, our data reveal a possible inhibitory effect of MBL at high concentrations in monocyte-DC transition, which probably provides one way of regulating adaptive immune responses by strict regulation of DCs, making MBL a better prospect for controlling relevant pathological events such as autoimmune diseases.

[Platelet-rich Plasma Induces M2 Macrophage Polarization via Regulating AMPK Singling Pathway].

OBJECTIVE: To investigate the role of platelet-rich plasma (PRP) in inducing the M2 macrophage polarization via regulating AMPK singling pathway. METHODS: The expressions of M1 marker CD11c and M2 marker CD206 in macrophages of blank control group, LPS group, LPS+PRP group, and LPS+PRP+Compound C group were detected by flow cytometry. Western blot was used to observe the effects of PRP on the expression of AMPK-mTOR signaling pathway-related proteins at different times (12 h, 18 h and 24 h) after LPS treatment. RNA interference technology was used to silence the expression of AMPK in macrophages, and the expression of TGF-ß protein was subsequently examined by Western blot. RESULTS: LPS significantly reduced the expression of CD206 and increased the expression of CD11c (P <0.05). After the addition of PRP, the expression of CD206 was significantly increased (P <0.05), while the expression of CD11c was significantly decreased (P <0.05). Compared with LPS group, PRP treatment significantly increased the expressions of p-AMPK and p-ULK1 proteins at 12 h, 18 h and 24 h, while significantly decreased the expression of p-mTOR protein (P <0.05). After the addition of AMPK inhibitor Compound C, the expression of CD206 was significantly reduced (P <0.05) and the expression of CD11c was significantly increased compared with LPS+PRP group (P <0.05). After silencing the expression of AMPK in macrophages, the promotion effect of PRP on TGF-ß was significantly reduced (P <0.05). CONCLUSION: PRP can stimulate the transformation of macrophages to M2 type via AMPK signalling pathway.

TCF12 activates MAGT1 expression to regulate the malignant progression of pancreatic carcinoma cells.

As a highly malignant gastrointestinal tumor, pancreatic carcinoma (PC) has poor prognosis due to its low early diagnosis rate, advanced tumor resection and chemotherapy resistance. Magnesium transporter 1 (MAGT1) is a magnesium ion transporter located on the cell membrane, which shows promotive effects on biological behaviors of multiple tumor cells. The aim of the present study was to investigate the role of MAGT1 in the progression of PC and its potential molecular mechanism. Based on the Gene Expression Profiling Interactive Analysis website, MAGT1 was highly expressed in tissues from patients with PC and was associated with poor prognosis. In functional experiments, MAGT1 was highly expressed in PC cell lines. The Cell Counting Kit-8, gap closure and Transwell assays, and western blot analysis, were used to investigate the effects of MAGT1 overexpression or knockdown on the biological behaviors of PC cells. It was found that MAGT1 promoted the proliferation, migration and invasion of PC cells in vitro. According to the Encyclopedia of RNA Interactomes website, transcription factor 12 (TCF12) mRNA expression level was positively correlated with MAGT1 expression level in the tissues from patients with PC. Positive targeting regulation of MAGT1 by TCF12 was also confirmed using a dual-luciferase gene reporter assay and chromatin immunoprecipitation. In addition, knockdown of TCF12 expression inhibited the proliferation and migration of PC cells, but overexpression of MAGT1 expression partly reversed this. These results suggested that TCF12 could promote the proliferation, migration and invasion of PC cells by activating MAGT1 expression, which was associated with poor prognosis. These findings suggest that MAGT1 could be a promising biomarker for the occurrence, progression and prognosis of PC.

Development of nanosilver doped carboxymethyl chitosan-polyamideamine alginate composite dressing for wound treatment.

Nowadays, treatment to the infected wounds caused by bacterial even multi-resistant bacterial strains and subsequently complete skin regeneration remain a critical clinical challenge. Herein, a novel multi-functional platform (Alg/1.0Ag@CMC-PAMAM/PRP) was prepared as wound dressings by mixing platelet rich plasma (PRP) with the sodium alginate (Alg) based dressing containing nano silver (Ag)-doped carboxymethyl chitosan grafted polyamideamine (Ag@CMC-PAMAM) cationic polymers. The present dressings exhibited high swelling, suitable water vapor transmission rate (WVTR), and good mechanical properties and degradability, as well as sustained release of PRP. Besides, the component of Ag@CMC-PAMAM nanoparticles endow them with excellent antibacterial performance, while the incorporation of PRP promotes the effect of anti-inflammatory and angiogenesis by up-regulating relative activity factor expression of TGF-ß1, CD31 and α-SMA and down-regulating the inflammatory-relative genes including TNF-α, IL-6 and IL-1ß, all of which promote the closure of wound and produce a superior healing effect to the commercial Aquacel Ag group. This work indicates that the prepared Alg/1.0Ag@CMC-PAMAM/PRP wound dressing is a promising biomaterial with synergistic effect of antibacterial property and wound healing.

Preparation of biocompatible wound dressings with dual release of antibiotic and platelet-rich plasma for enhancing infected wound healing.

The ever-growing threats of bacterial infection and chronic wound healing have provoked an urgent need for novel antibacterial wound dressings. In this study, we developed a wound dressing for the treatment of infected wounds, which can reduce the inflammatory period (through the use of gentamycin sulfate (GS)) and enhance the granulation stage (through the addition of platelet-rich plasma (PRP)). Herein, the sustained antimicrobial CMC/GMs@GS/PRP wound dressings were developed by using gelatin microspheres (GMs) loading GS and PRP, covalent bonding to carboxymethyl chitosan (CMC). The prepared dressings exhibited high water uptake capability, appropriate porosity, excellent mechanical properties, sustain release of PRP and GS. Meanwhile, the wound dressing showed good biocompatibility and excellent antibacterial ability against Gram-negative and Gram-positive bacteria. Moreover, in vivo experiments further demonstrated that the prepared dressings could accelerate the healing process of E. coli and S. aureus-infected full-thickness wounds in vivo, reepithelialization, collagen deposition and angiogenesis. In addition, the treatment of CMC/GMs@GS/PRP wound dressing could reduce bacterial count, inhibit pro-inflammatory factors (TNF-α, IL-1ß and IL-6), and enhance anti-inflammatory factors (TGF-ß1). The findings of this study suggested that biocompatible wound dressings with dual release of GS and PRP have great potential in the treatment of chronic and infected wounds.

Preparation and Evaluation of Sustained Release Platelet-Rich Plasma-Loaded Gelatin Microspheres Using an Emulsion Method.

The management and treatment of chronic wounds or acute wounds remain a major challenge in modern medicine. The application of autologous platelet-rich plasma (PRP) has become a promising adjuvant therapy to promote wound healing. PRP is derived from centrifuged whole blood to extract concentrated platelets, and a large amount of cytokines and growth factors are released upon activation. These bioactive molecules can enhance angiogenesis and tissue regeneration. Herein, PRP-loaded gelatin microspheres were prepared by the emulsion cross-linking method. Scanning electron microscopy results showed that the prepared microspheres are completely spherical, with an average particle size of 15.95 ± 3.79 µm and having a uniform particle size. Among them, the surface of a single microsphere is smooth and has a microporous structure, which may be the main channel for drug diffusion. Results of drug release measurements show that the prepared microspheres can slowly release the vascular endothelial growth factor for more than 7 days. In vitro cell experiments show that the prepared microspheres can promote proliferation and migration of L929 mouse fibroblast cells. In summary, the prepared PRP-loaded gelatin microspheres with high and long-term activity can provide experimental and theoretical knowledge for the development of the clinical long-acting injectable formulations.

[Research on piezoelectric protein sensor array for rapid detecting HIV(1+2) antibody].

To research piezoelectric immunosensor array for rapid detecting HIV(1+2), piezoelectric immunosensor array matrix was designed. HIV(1+2)C1 antigen was immobilized onto the silver electrodes of quartz-crystal microbalance, which was modified with adsorption and cross-linked method. In the clean air flow and monitoring environment, standard quality control and clinical serum sample were detected. The linear range for the measurement of HIV(1+2) was 0.01-0.2 NCU/ml. The sensitivity, specificity and accuracy of HIV(1+2) piezoelectric protein sensor array were 91.7%, 93.3% and 92.7% respectively.

Preparation of ADM/PRP freeze-dried dressing and effect of mice full-thickness skin defect model.

Acellular dermal matrix (ADM) and platelet-rich plasma (PRP) have each been used in wound healing. There are a few reports on the application of ADM with PRP in skin full-thickness defect models. In this study, the microstructure of ADM/PRP freeze-dried dressing was observed by scanning electron microscope. The PRP, ADM and PRP/ADM samples were recorded by Fourier transform infrared (FTIR) spectrometer by the KBr methods. The concentration of growth factor was measured by enzyme-linked immunosorbent assay. Then, we made a mice full-thickness defect model and treated it with saline, PRP, ADM and PRP/ADM, respectively. The wound size was measured and calculated on days 3, 5, 7, 10 and 14. The obtained results demonstrated that the ADM/PRP freeze-dried dressing resulted in earlier collagen development. FTIR analysis confirmed the integration of ADM and PRP in the prepared PRP/ADM scaffold. The concentration of bFGF, EGF, TGF-ß1 and PDGF-BB in the ADM/PRP group was lower than that in the PRP group. The wound healing rate of the ADM/PRP group was significantly promoted. Furthermore, the ADM/PRP group had significant revascularization, rapid epithelialization and a well-differentiated epidermis. The collagen fibers were regularly arranged. Accordingly, ADM/PRP freeze-dried dressing promotes wound healing and can be used in the skin full-thickness wound.

Effects of a protective agent on freeze-dried platelet-rich plasma.

: Freeze-drying is an effective means of storing platelets. In this study, we investigated the effects of a protective agent on freeze-dried platelet-rich plasma (FD-PRP) after a 12-week preservation period. Platelet structure was measured by transmission electron microscopy (TEM), and the expression levels of procaspase activating compound (PAC)-1 and CD62P were measured by flow cytometry. The levels of transforming growth factor-beta (TGF-ß), platelet-derived growth factor (PDGF) and vascular endothelial growth factor (VEGF) were determined by ELISA. The effect of FD-PRP on cell proliferation was measured by cell counting. TEM revealed that most platelets were intact, and their internal structure was evident. The expression levels of the platelet activation marker CD62P in FD-PRP and fresh PRP were 36.83% ±â€Š8.21 and 35.47% ±â€Š4.11, respectively, without a significant difference (P > 0.05). The expression levels of PAC-1 in FD-PRP and fresh PRP were 3.23% ±â€Š0.49 and 2.83% ±â€Š0.44, respectively, without a significant difference (P > 0.05). Upon activation of FD-PRP and fresh PRP by thrombin, the levels of TGF-ß, PDGF and VEGF were not significantly decreased in FD-PRP. Moreover, FD-PRP promoted cell proliferation in a manner similar to that of fresh PRP. The protective agent maintained the biological activity of FD-PRP after a 12-week preservation period.

Enhanced healing activity of burn wound infection by a dextran-HA hydrogel enriched with sanguinarine.

Burn wounds are associated with a series of risks, such as infection and pathologic scar tissue formation, which significantly delay wound healing and lead to complications. In this study, we successfully fabricated a dextran-hyaluronic acid (Dex-HA) hydrogel enriched with sanguinarine (SA) incorporated into gelatin microspheres (GMs), which had high porosity, good swelling ratio, enhanced NIH-3T3 fibroblast cell proliferation, and sustained SA release profile. The in vitro degradation results indicate that the SA/GMs/Dex-HA hydrogel can be degraded. The in vitro antibacterial tests showed that the SA/GMs/Dex-HA hydrogel can inhibit methicillin-resistant Staphylococcus aureus (MRSA) and Escherichia coli (E. coli). We evaluated the wound-healing effects and antibacterial properties of SA/GMs/Dex-HA hydrogels in a rat full-thickness burn infection model. The hematoxylin-eosin (H&E) and Masson's trichrome staining results of the SA/GMs/Dex-HA hydrogel showed that it improved re-epithelialization and enhanced extracellular matrix remodeling, and immunohistochemistry results showed that the expression of TGF-ß1 and TNF-α was decreased, while the TGF-ß3 expression was increased. Our findings suggest that the SA/GMs/Dex-HA hydrogel provides a potential way for infected burn treatment with high-quality and efficient scar inhibition.

High-activity chitosan/nano hydroxyapatite/zoledronic acid scaffolds for simultaneous tumor inhibition, bone repair and infection eradication.

Implanted biomaterials combined tumor inhibition and bone repair property are urgently needed to address the huge bone destruction and the high local recurrence following primary surgery in bone tumor therapy. In this work, a high-activity chitosan/nano hydroxyapatite (CS/nHA) scaffold containing zoledronic acid (CS/nHA/Zol) was prepared with a facile method. The prepared CS/nHA/Zol scaffolds exhibited excellent tumor inhibition property towards giant cell tumor of bone (GCT) in vitro through inducing cells apoptosis by up-regulating pro-apoptosis genes expression and reducing the osteoclastic activity of tumor cells by down-regulating osteoclastic genes. Meanwhile, the prepared scaffolds possessed well biocompatibility and osteoinductivity as compared to pure CS/nHA scaffolds. Furthermore, the prepared scaffolds also presented outstanding antibacterial activity against clinical pathogenic S. aureus and E. coli. These overall findings successfully demonstrated the prepared CS/nHA/Zol scaffolds had a multifunction of tumor therapy, bone repair, and antibacterium, which provides a new approach possessed promising advantages in bone tumor therapy.

Plasma Transfusion in Patients With Cirrhosis in China: A Retrospective Multicenter Cohort Study.

Patients with cirrhosis used to be associated with frequent use of blood components because of their complex disorder of hemostasis and bleeding complications. Recent findings have indicated that patients with cirrhosis have a state of "rebalanced" or even procoagulant hemostasis and have questioned the prophylactic use of plasma. To evaluate the current status of plasma use in patients with cirrhosis, we conducted a retrospective survey in 11 tertiary-care hospitals in China from September 1 to October 31, 2013. All patients admitted with cirrhosis during the study period were included in the study. The survey collected information including patients' diagnostic and demographic data, clinical course including bleeding complications and invasive procedures, laboratory results, and plasma transfusion data. Among 1595 patients with cirrhosis admitted to the 11 hospitals, 236 (14.8%) patients received 1 or more plasma transfusions during the study period. The number of plasma transfusions is defined as the number of transfusion orders. A total of 1037 plasma transfusions were administered to these patients, with a mean of 4.4 transfusions per transfused patient, ranging from 1 to 22 transfusions per transfused patient. Most plasma transfusions (760/1037; 73.3%) were given to patients without bleeding, for treatment of coagulopathy either without planned invasive procedures (70.4%) or before invasive procedures (2.9%). The median dose of plasma transfusion was 3.8 mL/kg. The rate of plasma transfusion of participating hospitals varied from 5.3% to 31.8%. It is encouraging to see that in one teaching hospital, 85.7% plasma transfusions were given to patients with bleeding indication, showing a promising sign in appropriate transfusion. Prophylaxis or empirical plasma transfusion is still a common problem in managing patients with liver cirrhosis. Wide variations are found in plasma transfusion practice among hospitals. Effective measures to control and reduce empirical correction of abnormal coagulation tests through transfusing plasma should be strengthened urgently.

A multicenter study of blood component transfusion in patients with liver cirrhosis in China: Patient characteristics, transfusion practice, and outcomes.

BACKGROUND: Cirrhosis is a complex acquired disorder of hemostasis and patients frequently receive blood transfusions. But there is very limited data on patterns of blood use at a patient level. AIMS: To characterize blood use in cirrhotic patients in China and compare with recommendations to help identify areas where quality improvement strategies can be targeted. We also compared findings to a similar study undertaken in UK. METHODS: A cross-sectional study was conducted in 11 hospitals over a 2-month period. Data were collected prospectively on each hospitalized cirrhotic patient to day 28. RESULTS: 1595 cirrhotic patients were included and 20.6% were transfused. 48.2% of transfused patients received transfusion for bleeding, most commonly gastrointestinal bleeding (65.8%). The remaining 51.8% were transfused for non-bleeding indications. 32.5% of patients transfused for gastrointestinal bleeding with red blood cells had a pre-transfusion haemoglobin >7g/dL. 89.1% of patients transfused frozen plasma for non-bleeding indications received them in the absence of a planned procedure. The patterns of blood transfusion in cirrhosis were different between China and UK. Of note, empirical prophylactic use of frozen plasma was more common in the Chinese study (89%) than in the UK (24%). CONCLUSION: Education and research should be implemented to improve patient blood management, especially in prophylactic frozen plasma use area.

Mannan binding lectin attenuates double-stranded RNA-mediated TLR3 activation and innate immunity.

Mannan binding lectin (MBL) functions as a pattern recognition molecule (PRM) which is able to initiate complement activation. Here, we characterize a previously unrecognized attribute of MBL as a double-stranded RNA (dsRNA) binding protein capable of modifying Toll like receptor 3 (TLR3) activation. MBL interacts with poly(I:C) and suppresses poly(I:C)-induced activation of TLR3 pathways and subsequent cytokine production. In addition, MBL binds to TLR3 directly. Surprisingly, disrupting the interaction between MBL and complement receptor 1 (CR1) or restraining the traffic of MBL to phagosome reversed the MBL limited TLR3 activation. We demonstrate the importance of MBL guided ligands intracellular localization, emphasizing the significance of understanding the dynamics of TLR agonists complexed with MBL or other PRMs inside the cell in immune defense.

Evaluation of the effects of homologous platelet gel on healing lower extremity wounds in patients with diabetes.

The treatment of chronic diabetic wounds remains complicated, despite new insight into the cellular and molecular basis of wound healing and cutaneous regeneration. A growing body of clinical trials has shown that platelet release has a notable effectiveness on refractory ulcer healing. However, patients with chronic diabetic ulcers usually have poor general health, and the large-volume blood absence required to produce autologous platelet-rich plasma often causes adverse effects. To overcome the limitation, the homologous platelet gel (PG) from healthy donor was used for the treatment of chronic diabetic lower extremity wound in the study. We show here that homologous derived platelets significantly enhanced EVC304 cell and HaCaT cell proliferation and homologous PG was capable of prompting cell migration. Twenty-one patients with refractory diabetic lower extremity ulcers, who had no response to conventional treatments, were treated in this study. Our data indicated that homologous PG was effective for the enhancement and acceleration of diabetic lower extremity wounds healing. We propose that homologous PG appeared to enhance vascularization and epithelialization, which might induce a quicker healing process and and encourage controlled studies in future.

Mannose-binding lectin inhibits monocyte proliferation through transforming growth factor-ß1 and p38 signaling pathways.

Mannose-binding lectin (MBL), a plasma C-type lectin, plays an important role in innate immunity. However, the interaction, and the consequences of it, between MBL and the immune system remain ill defined. We have investigated the contributing mechanisms and effects of MBL on the proliferation of human monocytes. At lower concentrations (≤4 µg/ml) MBL was shown to partially enhance monocyte proliferation. By contrast, at higher concentrations (8-20 µg/ml) of MBL, cell proliferation was markedly attenuated. MBL-induced growth inhibition was associated with G0/G1 arrest, down-regulation of cyclin D1/D3, cyclin-dependent kinase (Cdk) 2/Cdk4 and up-regulation of the Cdk inhibitory protein Cip1/p21. Additionally, MBL induced apoptosis, and did so through caspase-3 activation and poly ADP-ribose polymerase (PARP) cleavage. Moreover, transforming growth factor (TGF)-ß1 levels increased in the supernatants of MBL-stimulated monocyte cultures. We also found that MBL-dependent inhibition of monocyte proliferation could be reversed by the TGF-ß receptor antagonist SB-431542, or by anti-TGF-ß1 antibody, or by the mitogen-activated protein kinase (MAPK) inhibitors specific for p38 (SB203580), but not ERK (U0126) or JNK (SP600125). Thus, at high concentrations, MBL can affect the immune system by inhibiting monocyte proliferation, which suggests that MBL may exhibit anti-inflammatory effects.

The efficacy of autologous platelet-rich plasma combined with erbium fractional laser therapy for facial acne scars or acne.

The aim of this study was to evaluate the efficacy of autologous platelet-rich plasma (PRP) combined with erbium fractional laser therapy for facial acne or acne scars. PRP combined with erbium fractional laser therapy was used for the treatment of 22 patients, including 16 patients who suffered from facial acne scars and 6 patients who suffered from acne scars concomitant with acne. Whole blood (40 ml) was collected from each patient, and following differential centrifugation, PRP was harvested. After using an erbium fractional laser, we applied PRP to the entire face of every patient. Digital photos were taken before and after the treatment for evaluation by dermatologists and the patients rated the efficacy on a 5-point scale. The erythema was moderate or mild, while its total duration was <3 days; after receiving the treatment three times, 90.9% of the patients showed an improvement of >50%, and 91% of the patients were satisfied; no acne inflammation was observed after treatment. PRP combined with erbium fractional laser therapy is an effective and safe approach for treating acne scars or acne, with minimal side-effects, and it simultaneously enhanced the recovery of laser-damaged skin.