Uncertainty and influencing factors of the placement error of three-dimensional conformal intensity-modulated radiotherapy for thoracic tumors.

OBJECTIVE: Radiation therapy is an important method for the treatment of chest tumors. This study discussed the placement error of three-dimensional (3D) conformal intensity-modulated radiotherapy in patients with different types of chest tumors and analyzed the relevant influencing factors. PATIENTS AND METHODS: 100 patients with chest tumors diagnosed and treated in our hospital from March 2016 to March 2018 were randomly selected as research subjects, including 42 cases of esophageal cancer, 44 cases of breast cancer, and 14 cases of lung cancer. All patients underwent 3D conformal radiotherapy. The setup errors of patients with esophageal cancer, breast cancer, and lung cancer were detected after 3D conformal radiotherapy. Besides, the influencing factors of 3D conformal for thoracic tumors were analyzed by multiple linear regression analysis. RESULTS: After 3D conformal radiotherapy, the systematic errors of patients with esophageal cancer in X-axis, Y-axis, and Z-axis were -0.10, 1.26 and 0.07, respectively, while the random errors in X-axis, Y-axis, and Z-axis were 1.18, -1.14, and 0.97 respectively. The times for the absolute values of the positioning error with a range of ≤5 mm in X-axis, Y-axis, and Z-axis were 40 (95.24%), 2 (4.76%) and 36 (85.71%), while these with a range of >5 mm in X-axis, Y-axis, and Z-axis were 6 (14.29%), 41 (97.62%) and 1 (2.38%), respectively. For patients with breast cancer, the systematic errors and random errors in X-axis, Y-axis, and Z-axis are -0.19, 1.19, and 0.15, as well as 0.97, 0.02 and 1.29, respectively. The times for the absolute values of the positioning error with a range of ≤5 mm and >5 mm were 41 (93.18%), 3 (6.82%), and 36 (81.82%), as well as 8 (18.18%), 42 (95.45%) and 2 (4.55%), severally. For patients with lung cancer, the systematic errors and random errors in X-axis, Y-axis, and Z-axis were 0.14, 1.42, and 0.15, as well as 1.35, -0.23 and 1.12, respectively. The times for the absolute values of the positioning error with the range of ≤5 mm and >5 mm were 14 (93.33%), 1 (6.67%), and 11 (73.33%), as well as 4 (26.67%), 14 (93.33%) and 1 (6.67%) after 3D conformal radiotherapy. After multiple linear regression analyses, gender and lung volume were the influencing factors of Z-axis setup error, and the lesion location was the influence factor of Y-axis setup error (p<0.05). CONCLUSIONS: There are certain positioning errors in the X-axis, Y-axis, and Z-axis directions of thoracic tumors receiving 3D conformal radiotherapy. Gender, lung volume, and lesion location are all important factors that affect the placement error. The results of this study provide a certain reference for the positioning error of radiation therapy for thoracic tumors, which is conducive to improving the accuracy of radiotherapy and better protecting the surrounding tissues.

Setup error of electronic portal image device in IMRT for thoracic tumors and its influence.

OBJECTIVE: The aim of this study was to analyze the setup error of the electronics portal image device (EPID) in intensity-modulated radiation therapy (IMRT) for thoracic tumors and the influence on the outward expansion distance of the target area. PATIENTS AND METHODS: A total of 202 patients with chest tumors admitted to our hospital from March 2016 to March 2018 were selected as the observation subjects. All patients were treated with IMRT. The original plan was developed based on the SM90 obtained by the planning target volume (PTV) expansion method, and the new plan was obtained by shifting the isocenter coordinates of the treatment plan according to the positioning error value obtained by EPID. Before the treatment, EPID scans were performed. The electronic radiation field images (ERIs) were registered with the digitally reconstructed radiographic images (DRRs) generated by the treatment planning system using the image registration software, and the setup errors in the X, Y, and Z directions were further measured. The PTV was developed according to ERIs, and the setup error was simulated to obtain the PTV with 95% internal target volume (ITV) reaching the prescribed dose under the condition of a setup error. The outward expansion distance of clinical target volume (CTV) → PTV was calculated. RESULTS: In this experiment, the setup errors in X, Y, and Z directions were (-2.00±1.16) mm, (0.16±1.14) mm, and (-0.55±1.16) mm, respectively. The systematic error in the Z direction was -3.00 mm, and the random error in the X direction was 3.30 mm. The CTV → PTV outward expansion distance was set as 7, 8 and 7 mm in the X direction, Y direction and Z direction, respectively. At this time, under the presence of setup error, the PTV D95 and the ITV V100 in the new plan were (62.23±3.85) Gy and (97.51±1.56) %, respectively, effectively ensuring that 95% ITV of 90% patients reached the prescribed dose. In contrast, the ITV D95 and ITV V100 in the presence of setup error were (56.11±5.26) Gy and (90.15±3.12) %, respectively, at a CTV → PTV outward expansion distance of 5 mm, which could not guarantee that 95% ITV of 90% patients reached the prescribed dose. In the presence of a setup error, the double-lung 5 Gy irradiation of the total heart volume (V5), the double-lung 20 Gy irradiation of the total heart volume (V20), mean lung dose (MLD), mean heart dose (MHD), and D1 cm3 of the new plan increased by 0.89%, 0.29%, 0.13%, 0.06%, and 5 Gy, respectively, compared with the original plan. CONCLUSIONS: In general, the first treatment of radiotherapy in thoracic tumors mostly has a certain degree of setup error, which is most evident in the X direction. When the CTV → PTV outward expansion distance is set at 7, 8, and 7 mm in the X direction, Y direction, and Z direction, respectively, it can effectively ensure that 95% ITV reach the prescribed dose in 90% of patients in the presence of a setup error. EPID helps to achieve the desired effect of radiotherapy, improves the efficacy of radiotherapy, and reduces the side effects caused by radiotherapy errors.

The social and occupational consequences of the COVID-19 pandemic among patients with multiple sclerosis in three distinct populations: A web-based cross-sectional survey.

Background: The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes the disease COVID-19, has caused a pandemic that has rapidly affected the whole world and caused a significant threat to public health. The aim of this study was to investigate and analyze the social and occupational effects of the COVID-19 pandemic on patients with multiple sclerosis (MS) in three different countries: China, Spain, and Cuba. Methods: A cross-sectional survey was designed to assess the social and occupational effects of the COVID-19 pandemic in MS patients in these three countries, using a 25-item anonymous online questionnaire, structured into three sections. Quantitative data are expressed as mean (standard deviation), and quantitative data as absolute frequency and percentage. Results: A total of 361 participants responded to the questionnaire: 194 from China, 104 from Spain, and 63 from Cuba. We found no cases of COVID-19 among Chinese patients with MS, and few cases in Spain and Cuba. Respondents reported different levels of impact on relationships with friends, family, and colleagues, and patients in all three countries described increased use of digital or social media platforms. Spanish patients reported a significantly less negative impact than those in Cuba and China. Mental and cognitive effects were similar in all three countries, although China seemed to have a better situation. We also found that the time spent exercising decreased at specific points during the pandemic, but with few changes in dietary habits. Patients reported little or no change in their means of transport in all three countries. Most patients in all three countries reported little or no physical deterioration, especially in Chinese patients (82.47%), compared to the Spanish (70.20%) and Cuban respondents (73.02%). In general, patients from all three countries demonstrated confidence in overcoming the COVID-19 pandemic, although to a lesser extent among Spanish respondents. Conclusions: During the pandemic, family support was more effective in China than in Cuba and Spain. Neither COVID-19 infections nor the number of MS relapses increased significantly during lockdown in any of the three countries. Regarding their economic situation, Spanish MS patients reported a significantly less severe negative impact than those in Cuba and China. Patients from all three countries used digital or social media platforms more frequently, probably to maintain personal relationships. Chinese and Cuban respondents were more confident of the control of the pandemic than the Spanish, who were more pessimistic.

Introducción: El nuevo coronavirus de tipo 2, causante del síndrome respiratorio agudo severo o COVID-19, se ha expandido rápidamente a nivel mundial, convirtiéndose en una grave amenaza para la salud pública en forma de pandemia. El objetivo de este estudio es analizar los efectos sociolaborales de la pandemia de COVID-19 en pacientes con esclerosis múltiple (EM) en 3 países diferentes (China, España y Cuba). Métodos: Diseñamos un estudio transversal para valorar los efectos sociolaborales de la pandemia de COVID-19 en pacientes con EM procedentes de China, España y Cuba mediante un cuestionario digital de 25 preguntas divididas en 3 apartados. Los datos cuantitativos se expresan como medias y desviaciones estándar, mientras que los datos cualitativos se expresan mediante valores y porcentajes. Resultados: Un total de 361 pacientes respondieron al cuestionario (194 de China, 104 de España y 63 de Cuba). No encontramos ningún paciente chino con EM que hubiera padecido COVID-19, y los casos diagnosticados en España y Cuba fueron muy infrecuentes. A raíz de la pandemia, se observaron cambios en las relaciones con amigos, familiares y compañeros; además, los pacientes con EM usaron plataformas digitales y redes sociales con más frecuencia en los 3 países. El impacto negativo fue significativamente menor en España que en Cuba o China. Los efectos mentales y cognitivos de la pandemia fueron similares en los 3 países, aunque parece que la situación previa era mejor en China. Igualmente, observamos que el tiempo dedicado al ejercicio se redujo en momentos específicos durante la pandemia. Por el contrario, no se detectaron grandes cambios en los hábitos alimentarios. Los pacientes de los 3 países consideraron que no se produjeron cambios en su medio de transporte, o que estos fueron escasos. La mayoría de los pacientes ha experimentado poco o ningún deterioro físico, particularmente los pacientes chinos (82,47%) en comparación con los españoles (70,20%) y cubanos (73,02%). En líneas generales, todos los pacientes se mostraron esperanzados en superar la pandemia, aunque los pacientes españoles en menor grado. Conclusiones: Durante la pandemia, el apoyo familiar fue más importante en China que en Cuba o España. En ninguno de los 3 países se observó un aumento significativo en el número de casos de COVID-19 ni de brotes o recaídas durante el periodo de confinamiento. En términos económicos, los pacientes con EM españoles sufrieron un impacto negativo significativamente menor que los cubanos y chinos. Todos los pacientes usaron plataformas digitales o redes sociales con más frecuencia, probablemente con el fin de mantener relaciones personales. Los pacientes chinos y cubanos mostraron mayor confianza en el control de la pandemia que los españoles, que resultaron ser más pesimistas.

Three-dimensional digital reconstruction of skin epidermis and dermis.

This study describes how three-dimensional (3D) human skin tissue is reconstructed, and provides digital anatomical data for the physiological structure of human skin tissue based on large-scale thin serial sections. Human skin samples embedded in paraffin were cut serially into thin sections and then stained with hematoxylin-eosin. Images of serial sections obtained from lighting microscopy were scanned and aligned by the scale-invariant feature transform algorithm. 3D reconstruction of the skin tissue was generated using Mimics software. Fibre content, porosity, average pore diameter and specific surface area of dermis were analysed using the ImageJ analysis system. The root mean square error and mutual information based on the scale-invariant feature transform algorithm registration were significantly greater than those based on the manual registration. Fibre distribution gradually decreased from top to bottom; while porosity showed an opposite trend with irregular average pore diameter distribution. A specific surface area of the dermis showed a 'V' shape trend. Our data suggested that 3D reconstruction of human skin tissue based on large-scale serial sections could be a valuable tool for providing a highly accurate histological structure for analysis of skin tissue. Moreover, this technology could be utilized to produce tissue-engineered skin via a 3D bioprinter in the future.

L6H21 prolonged rats survival after limb allotransplantation by inhibiting acute rejection.

OBJECTIVE: Preventing and reducing allograft rejection play a far more important role in limb allotransplantation. We previously found L6H21 could inhibit LPS-induced (lipopolysaccharide LPS) overexpression inflammatory factors in macrophages and specifically targets to MD-2 (myeloid differential protein-2 MD-2) required for TLR4 (Toll-like receptor 4 TLR4) activation and represented an important therapeutic target in inflammatory disorders. Therefore, we evaluated the effect and explored the mechanism of L6H21 in rats' limb allograft model. MATERIALS AND METHODS: The efficacy of L6H21 was evaluated in limb allograft rats and cyclosporine (CY-A) was used as a positive control agent. T-Lymphocyte in blood was analyzed and dendritic cells (DCs) separated from spleens using flow cytometry. ELISA was used to measure serum cytokine levels. Analysis of protein expressions was performed using Western blotting. RESULTS: L6H21 reduced the risk of acute rejection and prolonged survival of limb allograft rats. At 3 d and 5 d post-transplant, the ratio of CD4+/CD8+ was decreased in L6H21 group. L6H21 suppressed the content of IL-1α at 7d, IL-5 and IL-10 at both 3 d and 7 d after transplantation. L6H21 decreased the protein expressions of IRF3, p-IRF3, P38, p-P38 and p-IκBα while increased IκBα expression and decreased the ratio of p-IRF3/ IRF3, p-P38/ P38, p-IκBα/IκBα correspondingly. CONCLUSIONS: L6H21 could reduce the risk of acute rejection and prolong the survival of limb allograft rats through inhibiting the ratio of CD4+/CD8+ in blood and serum cytokine levels and suppressing protein expressions of IRF3, p-IRF3, P38, p-P38 and p-IκBα in DCs. So, it may serve as a potential candidate for the treatment of allograft rejection.

Characterization of microparticles in patients with venous malformations of the head and neck.

OBJECTIVES: To determine the basic biochemical features of microparticles (MP) in patients with venous malformation (VM) of the head and neck. METHODS: Microparticles were isolated from peripheral venous blood of VM patients or healthy subjects and from lesional fluid of VM patients. Flow cytometry and real-time polymerase chain reaction were employed to determine the concentration, cellular origin, and RNA expression of obtained MP. A functional coagulation test was applied to measure the coagulant activity of MP. RESULTS: Circulating levels of total MP, platelet-derived MP, and endothelial MP were significantly elevated in VM patients and were consistently increased in VM patients with more extensive lesions. Lesional MP (MP from lesional fluid of VM) in VM patients were more abundant than circulating MP from VM patients or healthy subjects. Moreover, MP from VM patients displayed markedly distinct mRNA and microRNA expression compared with healthy subjects. Furthermore, MP from VM patients exhibited enhanced procoagulant activity, as evidenced by significantly shorter coagulation time. CONCLUSIONS: This study demonstrates for the first time that patients with VM have an altered MP profile and MP may be associated with VM-associated thrombogenesis. Further studies are required to explore the precise pathophysiological roles of MP in VM.

Endophilin B1 regulates EGFR endocytic degradation in prostate cancer cell.

Prostate cancer (Pca) is one of the most common types of cancer for elder men. Aberrant expression of epidermal growth factor receptor (EGFR) and EGFR downstream signaling have been known to contribute to disease progression in prostate cancer. EGF-stimulated EGFR is internalized and the process of endocytic degradation of EGFR mediates its signaling which is frequently dysregulated in many kinds of cancer. In the present study, we demonstrated that endophilin B1 expression was inhibited and EGFR expression was significantly increased in prostate cancer cell lines. We demonstrated that suppression of endophilin B1 increased EGFR levels via delaying EGFR internalization triggered by EGF and its intracellular degradation. Endophilin B1 decreased also sustained EGFR downstream signaling such as Erk1/2 phosphorylation in response to EGF stimulation and promoted prostate cancer cell proliferation which is EGF independent. Our data indicated that endophilin B1 mediated the biological function of EGFR in cancer cell proliferation through regulating the EGFR endocytic trafficking and downstream signaling.

Down-Regulation of Donor Kupffer Cell B7 Expression Reduced Recipient Lymphocyte Activation and Secretion of Interleukin-2 In Vitro.

BACKGROUND: Kupffer cell (KC), a kind of important antigen-presenting cell in liver, play an important role in the process of acute rejection after liver transplantation. The aim of this study was to investigate effect of suppression of donor KC B7 expression on recipient lymphocyte activation and secretion of interleukin-2 (IL-2) in vitro. METHODS: Liver ex vivo perfusion with collagenase IV and density-gradient centrifugation were used to isolate donor Lewis rat KCs. The interference fragments of the B7 molecule were designed to construct RNA interference vector pSilencer 3.1H1-Neo-B7 that was transfected into KCs of donor rat. Reverse-transcription polymerase chain reaction was used to detect the changes in the expression of B7 molecules in KCs. The transfected KCs were divided into 3 groups: A, control group; B, empty vector group; and C, RNA interference group. The lymphocytes of recipient Brown Norway (BN) rats were isolated and cocultured with the cells in the 3 groups. Enzyme-linked immunosorbent assay was used to detect the content of IL-2 in the culture supernate. Methylthiazolyl tetrazolium assay was used to detect the proliferation of lymphocytes. RESULTS: The yield rate of KCs was 5 × 10(7), and the cell viability was >98%. RNA interference vector had been successfully constructed and identified by means of enzyme digestion and sequencing. The expression of B7 in KCs decreased by 22% after RNA interference (P < .01). After coculturing with lymphocytes of BN rats, compared with the control group, the decreased expression of B7 significantly inhibited the activation and proliferation of lymphocytes as well as the secretion of IL-2 by lymphocytes. The proliferation of lymphocytes in recipient BN rats decreased by 49% (P < .01), and the secretion of IL-2 in the culture supernate decreased by 67% (P < .01). CONCLUSIONS: This study successfully constructed a B7 RNA interference vector, and applied it to assessing reduction of B7 expression in donor KCs. RNA interference significantly suppressed the activation of recipient T lymphocytes and secretion of IL-2 via the CD28/B7 costimulatory pathway and may induce immune tolerance in liver transplants.

High titer and yield ethanol production from undetoxified whole slurry of Douglas-fir forest residue using pH profiling in SPORL.

BACKGROUND: Forest residue is one of the most cost-effective feedstock for biofuel production. It has relatively high bulk density and can be harvested year round, advantageous for reducing transportation cost and eliminating onsite storage. However, forest residues, especially those from softwood species, are highly recalcitrant to biochemical conversion. A severe pretreatment for removing this recalcitrance can result in increased sugar degradation to inhibitors and hence cause difficulties in fermentation at high solid loadings. Here, we presented high titer ethanol production from Douglas-fir forest residue without detoxification. The strong recalcitrance of the Douglas-fir residue was removed by sulfite pretreatment to overcome the recalcitrance of lignocelluloses (SPORL). Sugar degradation to inhibitors was substantially reduced using a novel approach of "pH profiling" by delaying acid application in pretreatment, which facilitated the simultaneous enzymatic saccharification and fermentation of undetoxified whole slurry at a solid loading of 21%. RESULTS: "pH profiling" reduced furan production by approximately 70% in using SPORL pretreating Douglas-fir forest residue (FS-10) comparing with the control run while without sacrificing enzymatic saccharification of the resultant substrate. pH profiling also reduced carbohydrate degradation. The improved carbohydrate yield in pretreated solids and reduced fermentation inhibitors with pH profiling resulted in a terminal ethanol titer of 48.9 ± 1.4 g/L and yield of 297 ± 9 L/tonne FS-10, which are substantially higher, i.e., by 27% in titer and by 38% in yield, than those of a control SPORL run without pH profiling. CONCLUSIONS: Economical and large-volume production of commodity biofuels requires the utilization of feedstocks with low value (therefore low cost) and sustainably producible in large quantities, such as forest residues. However, most existing pretreatment technologies cannot remove the strong recalcitrance of forest residues to produce practically fermentable high titer sugars. Here, we demonstrated a commercially scalable and efficient technology capable of removing the strong recalcitrant nature of forest residues using "pH profiling" together with "low temperature SPORL". The resultant pretreated whole slurry of a Douglas-fir forest residue using this technology can be easily processed at high solids of 21% without detoxification to achieve a high ethanol yield of 297 L/tonne at 48.9 g/L. Graphical AbstractGraphic table of content.

Randomized clinical trial of autologous skin cell suspension combined with skin grafting for chronic wounds.

BACKGROUND: Treatment of chronic wounds using traditional surgical procedures is challenging because of the low graft take rates. This study investigated the combination approach of split-thickness autografts with harvested skin cell suspension for chronic wound treatment. METHODS: This randomized clinical trial enrolled patients with chronic wounds between March 2012 and December 2013. Patients who were assigned randomly to the active treatment received a split-thickness autograft combined with harvested skin cell suspension. Control patients received the split-thickness autograft alone. The primary outcome was the rate of complete wound closure by postoperative day 28. Analysis was by intention to treat. Patients who achieved wound closure were followed up for a minimum of 6 months to evaluate the quality of healing. RESULTS: A total of 88 patients were included, 44 in each group. More patients achieved complete wound closure in the skin cell group than in the control group (41 versus 34 patients; P = 0·035). Complete wound closure was observed at a median of 14 (95 per cent c.i. 12·0 to 16·0) days in the skin cell group and 20 (15·7 to 24·3) days in the control group (P = 0·001). The skin cell group had significantly fewer complications (4 versus 11 patients; P = 0·047). The autografted sites displayed better physical attributes and a reduced tendency for wound recurrence in the skin cell group. CONCLUSION: Complementary split-thickness autologous skin grafting with autologous skin cells harvested using ReCell® (Avita Medical, Cambridge, UK) technology improved the healing rate of chronic wounds. REGISTRATION NUMBER: UMIN000011966 (http://www.umin.ac.jp/ctr).

Expression of insulin-like growth factor-1 receptor in keloid and hypertrophic scar.

BACKGROUND: Keloid and hypertrophic scar (HS) are two pathological forms of excessive dermal fibrosis, which are due to aberrant wound-healing responses. Accumulating evidence suggests that aberrant activity of growth factors and increased numbers of growth factor receptors play an important role in the formation of pathological scar. AIM: We examined the expression level of insulin-like growth factor-1 receptor (IGF-IR) in keloid, HS and normal skin. METHODS: IGF-IR expression was analyzed by immunohistochemistry, real-time PCR and western blotting on tissues and fibroblasts from 30 patients, comprising 10 patients with keloid and 20 with HS (10 with immature and 10 with mature HS), and from 10 age-matched and sex-matched healthy controls. RESULTS: Immunoreactivity to IGF-IR was found in dermal fibroblasts of keloid (90%), immature HS, (80%) and mature HS (30%), but not in normal skin. There was no statistically significant difference in immunoreactivity scores between keloid and immature HS, but there was a significant difference (P < 0.01) between mature and immature HS. Real-time PCR and western blot analysis confirmed that there was high expression of IGF-IR in keloid and immature HS fibroblasts, but not in mature HS or normal skin fibroblasts. IGF-IR was expressed in the overlying epidermis, and there was no significant difference between the groups. CONCLUSIONS: IGF-IR may be involved in the pathogenesis of keloid and HS. Given that IGF-IR are predominantly expressed on dermal fibroblasts, targeting of IGF-IR in fibroblasts may be of benefit to prevent scarring.

Expression of liver-targeting peptide modified recombinant human endostatin and preliminary study of its biological activities.

Recombinant human endostatin (rEndostatin or endostar) has been shown to inhibit endothelial cells proliferation, migration, and angiogenesis and exhibits a broad spectrum of activities against solid tumors. However, rEndostatin is easily degradable and evenly distributed to all tissues. Selectively delivering rEndostatin to the lesion site might be more potent. The circumsporozoite protein (CSP) coats the malarial sporozoite and targets the liver for infection; I-plus of N end of CSP could specifically bind to the liver. Based on this, we hypothesize the fusion protein with introducing the CSP I-plus sequence into rEndostatin (rES-CSP) of which not only targets the liver, but also inhibits endothelial cells proliferation, migration, and tube formation. Therefore, it selectively reduces angiogenesis of hepatocellular carcinoma (HCC) and improves the anti-HCC effect. In this study, we synthesized a novel rES-CSP fusion gene by SOE-PCR and expressed the fusion protein in Escherichia coli BL2l (DE3). The suitable conditions were optimized by an orthogonal test (L(25)(5)(4)). The yields were 12 mg/l culture medium following refolding and purification on nickel-nitrilotriacetic acid (Ni-NTA) metal affinity chromatography matrices. The purified rES-CSP is specifically targeted to the hepatocyte and inhibited the proliferation and migration of human umbilical vein endothelial cells (HUVECs) in a dose-dependent manner and showed potent antiangiogenic capability on HUVECs tube formation assay and chick embryo chorioallantoic membrane (CAM) assay. These results lay the foundation for the further study of its targeting and anti-HCC in vivo and provide a feasible and convenient approach to produce liver-targeting drugs for treatment of the liver diseases.

An antennae-enriched carboxylesterase from Spodoptera exigua displays degradation activity in both plant volatiles and female sex pheromones.

Carboxyl/cholinesterase (CCE) is a large gene family of diverse functions, but in insects its function with respect to catabolism of sex pheromone components and plant volatiles is not well understood. In the present study, we cloned and functionally characterized one putative odorant-degrading enzyme (ODE) of the CCE family, SexiCXE14, from Spodoptera exigua. The tissue-temporal expression pattern revealed that the mRNA level of SexiCXE14 is antennae-enriched, sex equivalent and peaks at 3 days after moth eclosion. Functional study using the recombinant enzyme determined that SexiCXE14 has high degrading activity (Vmax) to host plant volatiles, suggesting its role in degradation of these volatiles. In addition, SexiCXE14 may also play a role in the degradation of sex pheromone components, as the Vmax and affinity parameter (Km) values with the sex pheromones are similar to those of reported pheromone degrading enzymes (PDEs). Further analysis of the relationship between substrate structure and enzymatic activity demonstrated that carbon chain length is a major influential factor, while the number of double bonds also affects the enzymatic activity. In addition, SexiCXE14 displays lower activity at acidic pH levels (pH 5.0) than in neutral conditions (pH 6.5). By characterizing this new ODE the present study provides insights in understanding of the high sensitivity of the moth olfactory system.

Downregulation of the transforming growth factor-ß/connective tissue growth factor 2 signalling pathway in venous malformations: its target potential for sclerotherapy.

BACKGROUND: Previous studies have implicated vascular destabilization and changes in extracellular matrix (ECM) composition in venous malformations (VMs). OBJECTIVES: To evaluate the expression levels of the connective tissue growth factor (CCN) family of matricellular proteins in VMs and explore their association with vascular destabilization. METHODS: The expression levels of CCNs 1-6, transforming growth factor (TGF)-ß, phosphorylated Tie2 and phosphorylated platelet-derived growth factor receptor ß in normal human skin tissues and VMs were detected by immunohistochemistry. Correlation between tested proteins was explored using the Spearman rank correlation test, followed by clustering analysis. In vitro studies using human umbilical vein endothelial cells (HUVECs) were performed for mechanism investigation. RESULTS: Expression of CCN2 was found to be strongly positive in fibroblast-like cells, endothelial cells and around blood vessels in normal human skin tissues, but it was significantly downregulated in VMs. Correlation analyses showed that expression levels of CCN2 and TGF-ß in VMs were positively correlated. The immunoreactivity of CCN2 was also closely correlated with perivascular α-smooth muscle cell actin(+) cell coverage in VMs. Moreover, in vitro studies in HUVECs indicated that CCN2 might act as a downstream target of TGF-ß, as demonstrated by the findings that treatment with exogenous TGF-ß or exogenous CCN2 could significantly upregulate the expression level of CCN2, and increase the expression levels of ECM components. Upregulation of the TGF-ß/CCN2 pathway was also detected in bleomycin-treated VM specimens. CONCLUSIONS: This study unmasks the downregulation of the TGF-ß/CCN2 pathway in VMs, and indicates its target potential for sclerotherapy.

Lignosulfonate-mediated cellulase adsorption: enhanced enzymatic saccharification of lignocellulose through weakening nonproductive binding to lignin.

BACKGROUND: Thermochemical pretreatment of lignocellulose is crucial to bioconversion in the fields of biorefinery and biofuels. However, the enzyme inhibitors in pretreatment hydrolysate make solid substrate washing and hydrolysate detoxification indispensable prior to enzymatic hydrolysis. Sulfite pretreatment to overcome recalcitrance of lignocelluloses (SPORL) is a relatively new process, but has demonstrated robust performance for sugar and biofuel production from woody biomass in terms of yield and energy efficiency. This study demonstrated the advantage of SPORL pretreatment whereby the presentation of lignosulfonate (LS) renders the hydrolysate non-inhibitory to cellulase (Cel) due to the formation of lignosulfonate-cellulase complexes (LCCs) which can mediate the Cel adsorption between lignin and cellulose, contrary to the conventional belief that pretreatment hydrolysate inhibits the enzymatic hydrolysis unless detoxified. RESULTS: Particular emphasis was made on the formation mechanisms and stability phase of LCCs, the electrostatic interaction between LCCs and lignin, and the redistributed Cel adsorption between lignin and cellulose. The study found that LS, the byproduct of SPORL pretreatment, behaves as a polyelectrolyte to form LCCs with Cel by associating to the oppositely charged groups of protein. Compared to Cel, the zeta potential of LCCs is more negative and adjustable by altering the molar ratio of LS to Cel, and thereby LCCs have the ability to mitigate the nonproductive binding of Cel to lignin because of the enlarged electrostatic repulsion. Experimental results showed that the benefit from the reduced nonproductive binding outweighed the detrimental effects from the inhibitors in pretreatment hydrolysate. Specifically, the glucan conversions of solid substrate from poplar and lodgepole pine were greatly elevated by 25.9% and 31.8%, respectively, with the complete addition of the corresponding hydrolysate. This contradicts the well-acknowledged concept in the fields of biofuels and biorefinery that the pretreatment hydrolysate is inhibitory to enzymes. CONCLUSIONS: The results reported in this study also suggest significant advantages of SPORL pretreatment in terms of water consumption and process integration, that is, it should abolish the steps of solid substrate washing and pretreatment hydrolysate detoxification for direct simultaneous saccharification and combined fermentation (SSCombF) of enzymatic and pretreatment hydrolysate, thereby facilitating bioprocess consolidation. Furthermore, this study not only has practical significance to biorefinery and bioenergy, but it also provides scientific importance to the molecular design of composite enzyme-polyelectrolyte systems, such as immobilized enzymes and enzyme activators, as well as to the design of enzyme separation processes using water-soluble polyelectrolytes.

Cyclosporine combined with nonlytic interleukin 2/Fc fusion protein improves immune response to hepatitis B vaccination in a mouse skin transplantation model.

BACKGROUND: Improving immune responses to vaccination in immunosuppressed patients is extremely important. Previously, we observed that cyclosporine (CsA) combined with a nonlytic interleukin (IL)-2/fragment crystallizable (Fc) fusion protein induces immune tolerance to mouse skin transplantations. In the present study, we asked whether this combination improved hepatitis B (HBV) vaccine efficacy in immunosuppressed mice while also prolonging skin graft survival. METHODS: After C57BL/6 mice received DBA/2 skin grafts, they were administered a 14-day course of CsA (30 mg/kg intraperitoneal) combined with IL-2/Fc (1 µg, intraperitoneal). HBV vaccine (2 µg) was injected intramuscularly on the day of skin transplantation. On day 14, the serum levels of hepatitis B surface antibody (HBsAb), IL-4, IL-10, interferon (IFN-γ), and IL-2 were measured by enzyme-linked immunosorbent assay. We assessed the percentages of CD4(+)CXCR5(+) follicular T helper cells, CD4(+)FoxP3(+) regulatory T cells (Treg) and expressions of IL-17, IL-21, FoxP3, Bcl-6 in the spleen. Animals were divided into four groups: control, vaccine-treated, CsA + vaccine-treated, CsA + IL-2/Fc + vaccine-treated hosts. RESULTS: Combination therapy significantly increased HBsAb levels and also prolonged skin graft survival. Serum levels of Th1 cytokines IL-2 and IFN-γ were significantly higher in the combination group, while Th2 cytokines, IL-4 and IL-10 were lower. Combined treatment increased the percentage of Treg and the expression of Foxp3 and IL-21, meanwhile inhibiting the expression of Bcl-6. But the percentage of Tfh did not significantly change among the groups. CONCLUSIONS: These observations suggested that a combination of CsA and IL-2/Fc fusion protein enhanced immune responses after HBV vaccination and prolonged skin graft survival.

Lignosulfonate and elevated pH can enhance enzymatic saccharification of lignocelluloses.

BACKGROUND: Nonspecific (nonproductive) binding (adsorption) of cellulase by lignin has been identified as a key barrier to reduce cellulase loading for economical sugar and biofuel production from lignocellulosic biomass. Sulfite Pretreatment to Overcome Recalcitrance of Lignocelluloses (SPORL) is a relatively new process, but demonstrated robust performance for sugar and biofuel production from woody biomass especially softwoods in terms of yields and energy efficiencies. This study demonstrated the role of lignin sulfonation in enhancing enzymatic saccharification of lignocelluloses - lignosulfonate from SPORL can improve enzymatic hydrolysis of lignocelluloses, contrary to the conventional belief that lignin inhibits enzymatic hydrolysis due to nonspecific binding of cellulase. RESULTS: The study found that lignosulfonate from SPORL pretreatment and from a commercial source inhibits enzymatic hydrolysis of pure cellulosic substrates at low concentrations due to nonspecific binding of cellulase. Surprisingly, the reduction in enzymatic saccharification efficiency of a lignocellulosic substrate was fully recovered as the concentrations of these two lignosulfonates increased. We hypothesize that lignosulfonate serves as a surfactant to enhance enzymatic hydrolysis at higher concentrations and that this enhancement offsets its inhibitive effect from nonspecific binding of cellulase, when lignosulfonate is applied to lignocellulosic solid substrates. Lignosulfonate can block nonspecific binding of cellulase by bound lignin on the solid substrates, in the same manner as a nonionic surfactant, to significantly enhance enzymatic saccharification. This enhancement is linearly proportional to the amount of lignosulfonate applied which is very important to practical applications. For a SPORL-pretreated lodgepole pine solid, 90% cellulose saccharification was achieved at cellulase loading of 13 FPU/g glucan with the application of its corresponding pretreatment hydrolysate coupled with increasing hydrolysis pH to above 5.5 compared with only 51% for the control run without lignosulfonate at pH 5.0. The pH-induced lignin surface modification at pH 5.5 further reduced nonspecific binding of cellulase by lignosulfonate. CONCLUSIONS: The results reported in this study suggest significant advantages for SPORL-pretreatment in terms of reducing water usage and enzyme dosage, and simplifying process integration, i.e., it should eliminate washing of SPORL solid fraction for direct simultaneous enzymatic saccharification and combined fermentation of enzymatic and pretreatment hydrolysates (SSCombF). Elevated pH 5.5 or higher, rather than the commonly believed optimal and widely practiced pH 4.8-5.0, should be used in conducting enzymatic saccharification of lignocelluloses.

Co-existent de novo myelodysplastic syndrome and T-cell non-Hodgkin lymphoma: a common origin or not?

The co-existence of de novo myelodysplastic syndrome (MDS) and non-Hodgkin lymphoma (NHL) prior to therapy is an extremely unusual finding. We report here a case of co-existent de novo MDS-refractory cytopenia with multilineage dysplasia and T-cell NHL, including clinical features, histopathological findings, molecular assessment, treatment course and outcomes. Other cases from the literature showing co-existence of both disorders are also reviewed; to date 19 similar cases have been reported. Among all cases (including the present patient), eight cases were diagnosed with de novo MDS and NHL simultaneously, which were considered to be true coincidences. The mechanisms responsible for the appearance of co-existence have not yet been ascertained, however in the present case a common chromosomal abnormality (20q deletion) was found in bone marrow and lymph node preparations. We conclude, therefore, that the co-existent de novo MDS and T-cell NHL seen in the present case may have a common origin.

Population pharmacokinetics of tacrolimus and CYP3A5, MDR1 and IL-10 polymorphisms in adult liver transplant patients.

BACKGROUND AND OBJECTIVE: Tacrolimus, an immunosuppressant widely used after liver transplantation, is characterized by a large inter-individual variability in its pharmacokinetics. The aim of this study was to perform population pharmacokinetic analysis of oral tacrolimus in liver transplant recipients and clarify the potential role of CYP3A5, MDR1 and IL-10 genetic polymorphisms in the variability of population pharmacokinetic parameters. METHODS: Tacrolimus blood concentration data (n = 1106) were collected from 104 full liver transplant patients and were analysed using a non-linear mixed-effects modelling program (nonmem). The CYP3A5*3, MDR1 G2677T/A and C3435T genetic polymorphisms were determined using polymerase chain reaction (PCR)-restriction fragment length polymorphism analysis. The IL-10 G-1082A variant was studied by allele-specific PCR method. RESULTS AND DISCUSSION: The liver function in patients as indicated by the total bilirubin level (TBIL) and different CYP3A5*3 genotypes in donors (CYPD) and recipients (CYPR) were observed to influence tacrolimus pharmacokinetic parameter of apparent clearance (Cl/F). The final regression model can be expressed as Cl/F = 15.9 - 1.88 TBIL + 7.65 CYPD + 7.00 CYPR. The relative standard errors (%RSE) of the parameter estimation were lower than 30% and the residual variability of tacrolimus trough blood concentration was 2.81 ng/mL. No significant effect of MDR1 and IL-10 polymorphisms was observed on population pharmacokinetic parameter of tacrolimus within 175 days after liver transplantation. CONCLUSION: The TBIL in patients and CYP3A5*3 genetic polymorphism in both donors and recipients contribute to the inter-individual variability of oral tacrolimus apparent clearance in Chinese adult liver transplant patients.

Transfection of CD40Ig into liver prevents acute rejection in rat liver transplantation.

OBJECTIVE: To investigate the effect of CD40Ig upon acute rejection of rat liver transplants. METHODS: Thirty-two orthotopic liver transplants were performed using Lewis to BN rats with "the two-cuff technique". The rats were randomly divided into three groups. Group A served as controls (n = 10); group B (n = 11) and group C (n = 11); Lipofectamine2000-pcDNA3.1 or Lipofectamine2000-pcDNA3.1. CD40Ig complex was injected into Lewis portal vein ex vivo before cold storage of the liver. On the fifth day after transplantation, three rats in each group were killed to study the pathological changes and TUNEL immune histochemistry performed to examine CD40Ig expression. Lymphocytes were obtained from the spleen. The mixed lymphocyte reaction (MLR) was performed to determine tolerance and sheep anti-human immunoglobulin G (IgG)-FITC-labeled T cells counted by flow cytometry. Postoperative survival times of rats in each group were recorded. The pathological changes of dead rats were observed. RESULTS: The mean survival times of group A and B were 11.00 +/- 4.28 and 12.75 +/- 5.57 days, respectively. There were serious acute rejections in allograft liver in groups A and B. Apoptosis index was 33.67 +/- 5.69 versus 39.00 +/- 5.29. Group C mean survival time was 41.25 +/- 13.70 days (P < .01). Immune histochemistry showed CD40Ig-positive elements in the allograft liver, which revealed light acute rejection and apoptosis index was 0.27 +/- 0.21 (P < .01). The part of the allografted liver in a dead rat showed light acute rejection while the others displayed chronic rejection. Recipients were specifically tolerant to donors in the MLR assay. The IgG-FITC-labeled T cells accounted for 11.57% of all T cells in group C. CONCLUSIONS: CD40Ig transfection inhibited T-cell costimulatory pathway, prevented acute rejection, and prolonged graft survival.