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Encouraging advances in both regenerative medicine and tissue engineering with stem cells require a short-term preservation protocol to provide enough time for quality control or the transportation of cell products from manufacturing facilities to clinical destinations. The hypothermic preservation of stem cells under refrigerated conditions (2-8 °C) in their specific culture medium provides an alternative and low-cost method for cryopreservation or commercial preservation fluid for short-term storage. However, most stem cells are vulnerable to hypothermia, which might result in cell damage from the cooling process and the lack of extracellular matrix (ECM). Herein, we report a peptide scaffold cell-culture-medium additive for mimicking

Background and ObjectivesSpontaneous spinal epidural hematoma (SSEH) is an uncommon but serious condition with a high morbidity rate. Although SSEH is related to numerous risk factors, its etiology remains unclear. There is a paucity of data on its prognostic factors. We aim to evaluate prognostic factors for SSEH in this study.MethodA retrospective study was performed on patients who were admitted for SSEH in 3 academic neurosurgical centers from January 2010 to June 2021. Clinical parameters, including clinical condition on admission, anticoagulants use, imaging modality, the timing and type of surgery performed, and outcomes, were collected. Prognostic factors were analyzed. The Frankel scale was used to assess the clinical condition.ResultsA total of 105 patients with SSEH were retrieved from medical records, with a mean age of 51.3 years. Eighty-three patients (79%) complained of acute onset of severe neck or back pain. Eighty-two patients (78%) suffered from moderate to severe neurologic deficits (Frankel scale A–C). Anticoagulation usage was found in 20% of cases. Lower thoracic spine (p = 0.046), use of anticoagulants (p = 0.019), sphincter function disfunction (p = 0.008), severe neurologic deficits at admission (p < 0.001), and rapid deterioration (p = 0.004) were found to be associated with poor outcomes. Surgical decompression was performed in 74 (70%) cases. The univariate and multivariate analysis revealed that preoperative severe neurologic deficits (p = 0.005) and extended paraplegia time (>12 hours, p = 0.004) were independent adverse prognostic factors. The univariate analysis revealed that lower thoracic spine location (p = 0.08) and rapid progression (p = 0.005) were correlated with poor prognosis, but the multivariate analysis failed to identify them as independent prognostic factors.DiscussionAdverse prognostic factors for SSEH might include thoracic segment location, use of anticoagulation, severe neurologic deficits on admission, sphincter dysfunction, and rapid progression. Preoperative neurologic deficit and extended paraplegia time were strongly correlated with the prognosis in the subset of patients who underwent surgical decompression. Timely surgical decompression is recommended for patients with moderate/severe neurologic deficits or progressive neurologic deterioration.

Human platelets (PLTs) are vulnerable to unfavorable conditions, and their adequate supply is limited by strict transportation conditions. We report here that PLTs preserved under three-dimensional (3D) conditions using novel biomimetic nanofiber peptides showed reduced apoptosis compared with classical PLTs stored at 22 °C and facilitated the storage and transportation of PLTs. The mechanism of PLT 3D preservation involves the formation of cross-links and a 3D nanofibrous network by a self-assembled peptide scaffold material at physiological conditions after initiation by triggers in plasma. PLTs adhere to the surface of the nanofibrous network to facilitate the 3D distribution of PLTs. The 3D microstructure, rheological properties, and effect on the inflammatory response and hemolysis were evaluated. Compared to traditional PLTs stored at 22 °C, PLTs subjected to 3D preservation showed similar morphology, number, aggregation activity, and reduced apoptosis. The detection of the reactive oxygen species (ROS) levels demonstrated that both reduced intracellular and mitochondrial ROS levels were correlated with reduced apoptosis. This study reveals a new 3D preservation method for PLTs based on the use of novel biomimetic nanofiber peptides that presents an attractive opportunity for various biomedical applications.

Mustards (Brassica juncea) are allopolyploid crops in the worldwide, and trichomes are essential quality attributes that significantly influence its taste and palpability in vegetable-use cultivars. As important accessory tissues from specialized epidermal cells, trichomes also play an important role in mitigating biotic and abiotic stresses. In this study, we constructed a F2 segregating population using YJ27 with intensive trichome leaves and 03B0307 with glabrous leaves as parents. By bulked segregant analysis (BSA-seq), we obtained a 2.1 Mb candidate region on B02 chromosome associated with the trichome or glabrous trait formation. Then we used 13 Kompetitive Allele Specific PCR (KASP) markers for fine mapping and finally narrowed down the candidate region to about 448 kb in length. Interestingly, among the region, there was a 3 kb sequence deletion that located on the BjuVB02G54610gene in the F2 individuals with trichome leaves. Genotyping results of F2 populations confirmed this deletion (R2=81.44%) as a major QTL. Natural population resequencing analysis and genotyping results further validated the key role of the 3 kb structure variation (SV) of insertion/deletion type in trichome development in B. juncea. Our findings provide important information on the formation of trichomes and potential target gene for breeding vegetable mustards.

Background Posttraumatic osteoarthritis (PTOA) correlates with a dramatic increase in multiple inflammatory factors after joint injury. There is a broad spectrum proteinase inhibitor known as alpha-2-macroglobulin (A2M) that plays an important role in protecting against inflammatory injury. Endogenous A2M is abundant in serum but insufficient in synovial fluid due to its large molecular weight and has a limited effect on articular cartilage inflammation. We hypothesize that intra-articular injection of A2M has a better clinical effect than the commonly used hyaluronic acid (HA) injection therapy. Furthermore, A2M could be a safer method than alternative treatments because it is a native substance in the human body. Method The in vivo effects of A2M and HA on cartilage degeneration were evaluated in rats with surgery-induced anterior cruciate ligament transection (ACLT) OA. One hundred rats were randomly divided into four groups (N = 25/group): (a) sham surgery + saline (Sham + S), (b) ACLT + A2M, (c) ACLT + HA, or (d) ACLT + saline (ACLT + S). Intra-articular injections of A2M were given immediately and 3 days after surgery, and then 20 µl was injected each weekly in each joint. The animals were sacrificed 12 weeks after surgery. Indian ink staining, safranin O staining and immunohistochemical staining were performed to assess cartilage damage. The extent of OA progression was graded by the Osteoarthritis Research Society International Osteoarthritis Cartilage Histopathology Assessment System (OOCHAS) (OA score = Grade × Stage; range, 0 to 24). The concentrations of matrix metalloproteinase-13 (MMP-13) and sGAG in synovial fluid lavage were measured by ELISA and spectrophotometric quantitative determination. OA-related gene expression was quantified by qPCR. Results Indian ink staining showed that the articular cartilage surface in rats treated with A2M was relatively intact compared with that in the animals treated with ACLT plus saline or HA injection, but there was no significant difference between the ACLT + HA group and the ACLT + S group. Histological staining indicated that early intra-articular injection of A2M attenuated OA pathogenesis in the rat ACLT model compared with that in animals treated with saline and HA. However, intra-articular injection of HA did not significantly protect cartilage against posttraumatic OA compared with saline treatment. The ELISA results showed that A2M reduced the concentration of MMP-13 in synovial fluid compared with that in the HA treatment group and other groups. RT–qPCR indicated that intra-articular A2M inhibited catabolism and enhanced anabolic metabolism, while there was no significant difference in the expression of OA-related genes between the ACLT + HA group and the ACLT + S group. Discussion In rat model, intra-articular injection of A2M had obvious protective effects against cartilage degeneration compared with HA treatment. The inflammatory factor MMP-13 provides strong evidence for this inhibitory effect. Moreover, we found no significant alleviation of articular cartilage pathogenesis in the HA-treated group, which suggests that the efficacy of HA is questionable and possibly transient, although it is still extensively used in clinical practice.

Human parvovirus B19 (B19V) and human parvovirus 4 (PARV4) are known to infect humans and transmit through contaminated blood and blood products. Globally, three genotypes of B19V, as well as PARV4, have been identified, respectively. The existence of different B19V genotypes in Chinese plasma donors has been investigated, however, the data regarding PARV4 were not available. The main objective of this study is to identify the genotypes of PARV4 circulating in Chinese plasma donors. By using a duplex quantitative polymerase chain reaction assay adapted for all genotypes of B19V and PARV4, 78 source plasma pools for fractionation were screened and quantified. Results showed that positive rates of B19V and PARV4 DNA in plasma pool samples were 25.64% and 14.10%, respectively. PARV4 sequences in two positive samples were next genotyped, and these two sequences belonged to PARV4 genotypes 1 and 2, respectively. In conclusion, the data present demonstrate the existence of PARV4 genotypes 1 and 2 in Chinese plasma donors for the first time and also show the relatively lower prevalence and level of PARV4 DNA in Chinese plasma donors in comparison with that of B19V DNA., Highlights At least two PARV4 genotypes, 1 and 2, were currently present in China. The prevalence and level of PARV4 DNA in Chinese plasma donors were relatively lower in comparison with that of B19V DNA. The rate of B19V and PARV4 coinfection in Chinese source plasma pools was low.

Background Dandy-Walker Syndrome (DWS) is a rare congenital brain malformation characterized by underdevelopment of cerebellar vermis and cystic enlargement of the fourth ventricle and enlargement of the posterior fossa. The cooccurrence of DWS and syringomyelia in adults is very rare. Case Description We report a man aged 19 years who presented with a 2-year history of tremor. Magnetic resonance imaging showed cystic dilation of the fourth ventricle, hypoplasia of the cerebellar vermis, and syringomyelia. Posterior fossa decompression and spinal cord ostomy were performed. Tremor was markedly improved and the fourth ventricular and the syringomyelia were reduced in size postoperatively. Conclusions Tremor can be a clinical manifestation in patients of DWS concomitant with syringomyelia in adults. Spinal cord ostomy combined with posterior fossa decompression may be an effective approach for alleviation of symptoms and syringomyelia.

Here we report a solution processed environmentally friendly MoS3 hole-transport material for Sb2Se3 solar cells, where MoS3 exhibits a matched energy level relative to Sb2Se3. In the synthesis, H2S produced by the thermal decomposition of (NH4)2MoS4 is found to efficiently eliminate the antimony oxide impurity formed on the Sb2Se3 surface. Finally, the all-inorganic Sb2Se3 solar cell delivers an efficiency of 6.86% with excellent stability.

Defects in a semiconductor dictate the carrier transport and recombination ability, which are critical factors that influence the power conversion efficiency in solar cells. In this study, we demonstrate that the introduction of tellurium is able to fine tune the atomic ratio of Se/Sb in Sb2Se3 thin films, both Se-rich and Sb-rich Sb2Se3 are obtained. Using fine-tuned device fabrication and deep level defect spectroscopy for characterization, we experimentally disclose that Se-rich Sb2Se3 favors the formation of SeSb and VSb defects, while Sb-rich films benefit the formation of SbSe and VSe defects. With an appropriate excess of Se in Sb2Se3, a net efficiency improvement of 2% is obtained when compared with pristine Sb2Se3 based solar cells. Our study provides an effective strategy to manipulate the defect formation in Sb2Se3 solar cells and inspires further improvement in the efficiency of Sb2Se3 solar cells.

Hull-less pumpkins (

Gene mining in a C. lanatus × C. amarus population revealed one gene, ACS7, linked to primary root elongation in watermelon. Watermelon is a xerophytic crop characterized by a long primary root and robust lateral roots. Therefore, watermelon serves as an excellent model for studying root elongation and development. However, the genetic mechanism underlying the primary root elongation in watermelon remains unknown. Herein, through bulk segregant analysis we identified a genetic locus, qPRL.Chr03, controlling primary root length (PRL) using two different watermelon species (Citrullus lanatus and Citrullus amarus) that differ in their root architecture. Fine mapping revealed that xaa-Pro dipeptidase and 1-aminocyclopropane-1-carboxylate synthase 7 (ACS7) are candidate regulators of the primary root growth. Allelic variation in the delimited region among 193 watermelon accessions indicated that the long-root alleles might only exist in C. amarus. Interestingly, the discrepancy in PRL among the C. amarus accessions was clearly associated with a nonsynonymous single nucleotide polymorphism variant within the ACS7 gene. The ACS7 expression and ethylene levels in the primary root tips suggested that ethylene is a negative regulator of root elongation in watermelon, as supported by the application of 1-aminocyclopropane-1-carboxylate (ACC, the ethylene precursor) or 2-aminoethoxyvinyl glycine (AVG, an ACS inhibitor). To the best of our knowledge, these findings provide the first description of the genetic basis of root elongation in watermelon. The detected markers of the ACS7 gene will facilitate marker-assisted selection for the PRL trait to improve water and nutrient use efficacy in watermelon and beyond.

The stromal vascular fraction (SVF) derived from adipose tissue contains heterogeneous cell populations and has enormous potential for clinical therapy. There are two main methods for SVF isolation: enzymatic isolation and mechanical isolation, both of which have shortcomings. In this study, optimized conditions for the isolation of high-quality SVF were established, and applications in fat grafting were evaluated.Adipose tissue was chopped into small pieces and then ground into an erosive shape using a syringe. The pieces were digested with 0.15% type II collagenase for 35 min at 37 °C. After centrifugation, the pellets were resuspended in DMEM and passed through a 100-μm strainer. The filtered cells were analyzed by flow cytometry. The fat graft was enriched with isolated SVF and subcutaneously transplanted into nude mice. Three weeks after transplantation, grafts were isolated, and HE staining, immunocytochemistry, and western blotting were conducted.The harvested SVF cells reached2 × 10Our study provides an efficient procedure for SVF isolation, its application in fat grafting, and possible underlying mechanisms.This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Background The pathogenesis of transfusion-related acute lung injury (TRALI) progress is incompletely understood, and specific therapies for TRALI are lacking. Alveolar macrophages (AMs) are critical for initiation and resolution of lung inflammation. However, the role of AMs in the pathogenesis of TRALI-associated lung failure is poorly understood. Study design and methods Mouse model for in vivo imaging of interleukin (IL)-6 activation in AMs was established by intratracheal instillation of a lentiviral vector carrying the luciferase reporter gene. The TRALI mouse model was produced by intraperitoneal lipopolysaccharide plus intravenous major histocompatibility complex Class I monoclonal antibody treatment. We focused on the changes in AMs in the lung during TRALI and examined whether targeting AMs is an effective strategy to alleviate this condition. Measurements and main results We confirmed that TRALI progress is accompanied by IL-6 activation in AMs. Further study showed that AMs undergo M1 activation during TRALI progress. AM depletion protected mice from TRALI, and transfusion of M1-polarized AMs into 34-1-2 s-treated mice elevated acute lung injury, indicating that the severity of TRALI was able to be ameliorated by targeting AM polarization. Next, we showed that α1 -antitrypsin (AAT) expression improved lung injury by modulating the production of IL-6 in AMs and decreased polarization of AMs toward the M1 phenotype. Conclusions M1-polarized AMs are crucial in a mouse model of TRALI, and AAT may serve as a future treatment for TRALI by regulating the polarization of AMs.

Antimony selenide (Sb2Se3) has attracted increasing attention in photovoltaic applications due to its unique quasi-one-dimensional crystal structure, suitable optical band gap with a high extinction coefficient, and excellent stability. As a promising light-harvesting material, the available synthetic methods for the fabrication of a high-quality film have been quite limited and seriously impeded both the fundamental study and the efficiency improvement. Here, we developed a facile and low-cost hydrothermal method for in situ deposition of Sb2Se3 films for solar cell applications. In this process, we apply KSbC4H4O7 and Na2SeSO3 as the antimony and selenium sources, respectively, in which thiourea (TU) serves as an additive to suppress the formation of Sb2O3 impurities. As a result, improved phase purity and enhanced crystallinity of the Sb2Se3 film are thus obtained, along with decreased trap states. Finally, the planar heterojunction Sb2Se3 solar cell delivered a power conversion efficiency of 7.9%, which is thus far the highest reported efficiency among solution-processed Sb2Se3 solar cells. This simple procedure and efficiency achievement demonstrate the great potential of the hydrothermal deposition process for the fabrication of high-efficiency Sb2Se3 solar cells.

Here we report a solution processed environmentally friendly MoS3 hole-transport material for Sb2Se3 solar cells, where MoS3 exhibits a matched energy level relative to Sb2Se3. In the synthesis, H2S produced by the thermal decomposition of (NH4)2MoS4 is found to efficiently eliminate the antimony oxide impurity formed on the Sb2Se3 surface. Finally, the all-inorganic Sb2Se3 solar cell delivers an efficiency of 6.86% with excellent stability.

In this study, we provide fundamental understanding on defect properties of the Sb

α1-antitrypsin (AAT) is a 52kDa serine protease inhibitor that is abundant in plasma. It is synthesized mainly by hepatic cells, and widely used to treat patients with emphysema due to congenital deficiency of AAT. A new isolation method for the purification of AAT from Cohn Fraction IV (Cohn F IV) is described. Cohn F IV is usually discarded as a byproduct from Cohn process. Using Cohn F IV as starting material does not interfere with the production of other plasma proteins and the cost of purification could be reduced greatly. Parameters of each step during purification were optimized, 15% polyethyleneglycol (PEG) concentration and pH 5.2 for PEG precipitation, elution with 0.05M sodium acetate and pH 4.7 for ion-exchange chromatography, and two steps blue sepharose affinity chromatography were chosen for AAT purification. The final protein with purity of 98.17%, specific activity of 3893.29 IU/mg, and yield of 28.35%, was achieved. Western blotting was applied for qualitative identification of final product, which specifically reacted with goat anti-human AAT antibody. LC-ESI-MS/MS was also employed to confirm the final protein. High performance liquid chromatography was used to analyze the composition of purified protein suggesting that pure protein was achieved. The molecular weight of AAT is 51062.77Da which was identified by LC-MS-MS. The manufacturing process described here may make better use of human plasma with Cohn F IV as starting material. The simple process described in this study is simple and inexpensive, it has a potential value for large scale production.

Summary Fruit rind plays a pivotal role in alleviating water loss and disease and particularly in cracking resistance as well as the transportability, storability and shelf‐life quality of the fruit. High susceptibility to cracking due to low rind hardness is largely responsible for severe annual yield losses of fresh fruits such as watermelon in the field and during the postharvest process. However, the candidate gene controlling the rind hardness phenotype remains unclear to date. Herein, we report, for the first time, an ethylene‐responsive transcription factor 4 (ClERF4) associated with variation in rind hardness via a combinatory genetic map with bulk segregant analysis (BSA). Strikingly, our fine‐mapping approach revealed an InDel of 11 bp and a neighbouring SNP in the ClERF4 gene on chromosome 10, conferring cracking resistance in F2 populations with variable rind hardness. Furthermore, the concomitant kompetitive/competitive allele‐specific PCR (KASP) genotyping data sets of 104 germplasm accessions strongly supported candidate ClERF4 as a causative gene associated with fruit rind hardness variability. In conclusion, our results provide new insight into the underlying mechanism controlling rind hardness, a desirable trait in fresh fruit. Moreover, the findings will further enable the molecular improvement of fruit cracking resistance in watermelon via precisely targeting the causative gene relevant to rind hardness, ClERF4.

Objective. We aimed to evaluate the efficacy of anterior approach microscopic surgery for patients with the pincer mechanism in cervical spondylotic myelopathy. Methods. The clinical data of pincer cervical spondylotic myelopathy that received anterior cervical decompression and fusion in our hospital from Aug 2014 to Dec 2017 were analyzed retrospectively, including 12 males and 9 females, with an average age of 64.3 years (range 46-81 years). Occupying rate, anterior occupying rate, and posterior occupying rate were measured on pre- and postoperative mid-sagittal MRIs. Pre- and postoperative Japanese Orthopedic Association (JOA) scores, intervertebral space height, and C2 to C7 Cobb’s angle were analyzed. Result. Duration of follow-up was six months. The pre- and postoperative anterior occupying rate were averagely 38.6±8.5% and 12.9±5.5%, respectively, the posterior occupying rates were averagely 27.4±7.2% and 13.1±6.6%, respectively, and Cobb’s angle changed from 15.3±8.0° to 22.7±7.9°. The intervertebral space height increased from 4.6±0.4mm to 6.5±0.4mm. JOA scores improved significantly by 59.4±34.0% at six months after surgery. Conclusion. Decompression by anterior microscopic surgery can increase spinal canal volume directly, recover intervertebral space height, and resize Cobb’s angle, but decrease the posterior compression by ligament Flava indirectly. Anterior decompression under the microscope may provide an alternative surgical option for partial patients with the pincer mechanism in cervical spondylotic myelopathy.

Antimony selenosulfide (Sb2 (S,Se)3 ) is an emerging low-cost, nontoxic solar material with suitable bandgap and high absorption coefficient. Developing effective methods for fabricating high-quality films would benefit the device efficiency improvement and deepen the fundamental understanding on the optoelectronic properties. Herein, equipment is developed that allows online introduction of precursor vapor during the reaction process, enabling sequential coevaporation of Sb2 Se3 and S powders for the deposition of Sb2 (S,Se)3 thin films. With this unique ability, it is revealed that the deposition sequence manipulates both the interfacial properties and optoelectronic properties of the absorber film. A power conversion efficiency of 8.0% is achieved, which is the largest value in vapor-deposition-derived Sb2 (S,Se)3 solar cells. The research demonstrates that multi-source sequential coevaporation is an efficient technique to fabricate high-efficiency Sb2 (S,Se)3 solar cells.

Human cytomegalovirus (hCMV), a ubiquitous beta-herpesvirus, has been associated with several autoimmune diseases. However, the direct role of hCMV in inducing autoimmune disorders remains unclear. Here we report the identification of an autoantibody that recognizes a group of peptides with a conserved motif matching the Pp150 protein of hCMV (anti-Pp150) and is shared among patients with various autoimmune diseases. Anti-Pp150 also recognizes the single-pass membrane protein CIP2A and induces the death of CD56(bright) NK cells, a natural killer cell subset whose expansion is correlated with autoimmune disease. Consistent with this finding, the percentage of circulating CD56(bright) NK cells is reduced in patients with several autoimmune diseases and negatively correlates with anti-Pp150 concentration. CD56(bright) NK cell death occurs via both antibody- and complement-dependent cytotoxicity. Our findings reveal that a shared hCMV-induced autoantibody is involved in the decrease of CD56(bright) NK cells and may thus contribute to the onset of autoimmune disorders.

In this study, we provide fundamental understanding on defect properties of the Sb2(S,Se)3 absorber film and the impact on transmission of photo-excited carriers in N–i–P architecture solar cells by both deep level transient spectroscopy (DLTS) and optical deep level transient spectroscopy (ODLTS) characterizations. Through conductance–voltage and temperature-dependent current–voltage characterization under a dark condition, we find that the Sb2(S,Se)3 solar cell demonstrates good rectification and high temperature tolerance. The DLTS results indicates that there are two types of deep level hole traps H1 and H2 with active energy of 0.52 eV and 0.76 eV in the Sb2(S,Se)3 film, and this defect property is further verified by ODLTS. The two traps hinder the transmission of minority carrier (hole) and pinning the Fermi level, which plays a negative role in the improvement of open-circuit voltage for Sb2(S,Se)3 solar cells. This research suggests a critical direction toward the efficiency improvement of Sb2(S,Se)3 solar cells.

Cellular therapies represent a new frontier in the treatment of neurological diseases. Accumulating evidence from preclinical studies of animal models suggests that mesenchymal stromal cells (MSCs), also known as mesenchymal stem cells, are an effective therapy for neurological diseases. In this study, we established human MSC lines from both cranial bone marrow (cBMMSCs) and iliac crest bone marrow (iBMMSCs) from the same donors and found that cBMMSCs show higher expression of neural crest-associated genes than iBMMSCs. Moreover, as observed in both mRNA and protein assays, neurogenic-induced cells from cBMMSCs expressed significantly higher levels of neural markers, such as NESTIN, SLUG, SOX9, and TWIST, than those from iBMMSCs. Thus, cBMMSCs showed a greater tendency than iBMMSCs to differentiate into neuron-like cells.

α1-Antitrypsin (AAT) is widely used to treat patients with congenital AAT deficiency. Cohn Fraction IV (Cohn F IV) is normally discarded during the manufacturing process of albumin but contains approximately 33% of plasma AAT. We established a new process for large-scale purification of AAT from it. liquid chromatography-electrospray ionization-tandem mass spectrometry and high-performance liquid chromatography were applied for qualitative identification and composition analysis, respectively. Stabilizers were optimized for AAT activity protection during lyophilization and dry-heat. Virus inactivation by dry-heat and solvent/detergent (S/D) was validated on a range of viruses. AAT with purity of 95.54%, specific activity of 3,938.5 IU/mg, and yield of 26.79%, was achieved. More than 95% activity was reserved after S/D. More than 96% activity was obtained after lyophilization or dry-heat. After S/D, pseudorabies virus (PRV) and vesicular stomatitis virus (VSV) were inactivated below detectable level within 1 H. Virus titer reductions of more than 5.50 log10 and 5.38 log10 were achieved for PRV and VSV, respectively. Porcine parvovirus and encephalomyocarditis virus were inactivated by 3.17 log10 and 5.88 log10 reduction after dry-heat. The advantages of this process, including suitability for large-scale production, high purity, better utilization of human plasma, viral safety, commercial and inexpensive chromatography medium, may facilitate its further application.

Objective: Porcine endogenous retroviruses (PERV) involved in pig to human xenotransplantation have raised great concerns because of their ubiquitous nature in pigs and their ability of infecting human cells in vitro. Although no significant cytopathic effect attributed to PERV was evident on PERV-infected human embryonic kidney 293 (HEK293) cells, we did proteomic analysis to investigate the differences of protein profile in order to further characterize the effect of PERV infection. Methods: HEK293 cells were cocultured with porcine peripheral blood mononuclear cells (PBMCs). Protein profiles of PERV-infected and -noninfected HEK293 cells were analyzed by two-dimensional gel electrophoresis (2-DE). Protein spots with at least 1.5-fold alteration were identified by high-definition mass spectrometry (HDMS) analysis. Then real-time RT-PCR and Western blotting were performed to validate the proteomic results. Results: Differential analysis of PERV-infected and -noninfected HEK293 cells by 2-DE revealed ten differentially regulated proteins. The proteins identified by HDMS were involved in various cellular pathways including signal transduction, cell apoptosis, and protein synthesis. Conclusion: The results of this study revealed differentially expressed proteins in HEK293 cells cocultured with porcine PBMCs and implied that these changes were probably induced by PERV infection. These results provide clues and potential links to understanding the molecular effect of the infection by human-tropic PERV.

Human parvovirus B19 (B19V) and human parvovirus 4 (PARV4) are two parvoviruses known to infect humans and transmit through blood and plasma derived medicinal products (PDMPs). Inactivation of the two parvoviruses has proven to be difficult and nucleic acid testing (NAT) would be an efficient means to exclude viruses. In this study, an internally controlled multiplex quantitative real-time PCR (qPCR) assay for B19V and PARV4 simultaneous detection and quantification was established and evaluated. The optimized multiplex qPCR assay allowed for simultaneous detection of all of the genotypes (1-3) of B19V and PARV4, with equal limit of quantification (LOQ) of 5 copies/μL, rather than other blood-borne viruses. It had a wide dynamic range of reliable amplification linearity of at least 8 orders of magnitude. Low standard deviations (SD) of quantification cycle (Cq) values and low coefficients of variation (CV) of copy numbers for both B19V and PARV4 suggested a high level of repeatability and reproducibility for the multiplex qPCR assay. This multiplex qPCR assay can be served as a readily applicable approach to screen plasma units intended for further manufacturing into PDMPs to reduce the risk of parvoviruses infection by such products and may also be useful for the detection of B19V/PARV4 co-infection or co-existence.

Human parvovirus B19 (B19V) is a common human pathogen which is associated with a broad spectrum of clinical manifestations. Since B19V can be transmitted via blood and plasma derivatives, it has been considered to pose great risk in transfusion safety. Since the early 21st century, European Pharmacopoeia, the Plasma Protein Therapeutics Association and U.S. Food and Drug Administration have proposed a list of guidelines to reduce the risk of B19V transmission by plasma derivatives. In terms of blood, some countries implement management measures on B19V to maximize the safety of blood transfusion. In China, no related documentation for monitoring B19V has been issued. The aim of this review is to discuss the risk for B19V transmission through blood and plasma derivatives in China. The issues raised with the intention to contribute to further development of risk management measures.

Background Human parvovirus B19 (B19V) is a frequent contaminant of blood and plasma-derived medicinal products. Three distinct genotypes of B19V have been identified. The distribution of the three B19V genotypes has been investigated in various regions or countries. However, in China, data on the existence of different B19V genotypes are limited. Methods One hundred and eighteen B19V-DNA positive source plasma pool samples collected from three Chinese blood products manufacturers were analyzed. The subgenomic NS1/VP1u region junction of B19V was amplified by nested PCR. These amplified products were then cloned and subsequently sequenced. For genotyping, their phylogenetic inferences were constructed based on the NS1/VP1-unique region. Then putative recombination events were analyzed and identified. Results Phylogenetic analysis of 118 B19V sequences attributed 61.86 % to genotype 1a, 10.17 % to genotype 1b, and 17.80 % to genotype 3b. All the genotype 3b sequences obtained in this study grouped as a specific, closely related cluster with B19V strain D91.1. Four 1a/3b recombinants and 5 new atypical B19V variants with no recombination events were identified. Conclusions There were at least 3 subtypes (1a, 1b and 3b) of B19V circulating in China. Furthermore, putative B19V 1a/3b recombinants and unclassified strains were identified as well. Such recombinant and unclassified strains may contribute to the genetic diversity of B19V and consequently complicate the B19V infection diagnosis and NAT screening. Further studies will be required to elucidate the biological significance of the recombinant and unclassified strains. Electronic supplementary material The online version of this article (doi:10.1186/s12985-016-0611-6) contains supplementary material, which is available to authorized users.

Pasteurization is regularly used to inactivate viruses for the safety of plasma derivatives. Influence of pasteurization at 60 °C for 10 h on α2-Macroglobulin activity and virus inactivation were studied. With 40% sugar as stabilizers more than 70% α2-Macroglobulin activity was reserved after pasteurization compared with 20% in control. Glucose presented a better activity protection effect than sucrose and maltose. By pasteurization without stabilizer the virus titers of pseudorabies virus, Sindbis virus, porcine parvovirus and encephalomyocarditis virus were reduced more than 5.88 log10, 7.50 log10, 4.88 log10, and 5.63 log10 respectively within 2 h. By pasteurization with 40% glucose vesicular stomatitis virus was inactivated more than 5.88 log10 within 1 h. Only 2.71 log10 reduction was achieved for encephalomyocarditis virus after 10 h. 40% glucose protected α2-M activity and viruses simultaneously from pasteurization. Other viral inactivation methods need to be incorporated to ensure viral safety of this manufacturing process of α2-Macroglobulin.

Background α2-Macroglobulin (α2-M) has a curative effect on radiation injury. Virus transmission through plasma derivatives is still not risk-free. Effect of dry heat on α2-M activity and virus inactivation by dry heat in a new manufacturing process of α2-M were studied. Study design and methods Effects of 100°C for 30 minutes, 80°C for 72 hours, and lyophilization on α2-M activity were detected, and stabilizing agents were optimized. Effect of a treatment at 100°C for 30 minutes has been tested on a range of viruses and characteristics change of α2-M was investigated. Results More than 90 and 80% α2-M activity recovery were reserved after treatment at 100°C for 30 minutes and 80°C for 72 hours, respectively. A concentration of 0.05 mol/L histidine presented a better protecting effect for α-M activity. No substantial changes were observed in the characteristics of α2-M compared with the untreated. By lyophilization and dry-heat treatment at 100°C for 30 minutes, murine encephalomyocarditis virus and pseudorabies virus (PRV) were inactivated below detectable level within 5 minutes (virus titers reduction ≥ 5.75 log) and 30 minutes (virus titers reduction ≥ 6.00 log), respectively. Bovine viral diarrhea virus and porcine parvovirus were inactivated by 4.29 and 2.46 log reduction, respectively. Conclusion Treatment at 100°C for 30 minutes could improve the virus safety of α2-M with a slight activity loss.

As an abundant plasma protein, α2-macroglobulin (α2-M) participates widely in physiological and pathological activities including coagulation regulation, antitumor activities, and regulation of cytokines. It also presents a therapeutic potential for radiation injury. A two-step isolation method for the purification of α2-M from Cohn Fraction IV is described. This process includes a salting-out method and immobilized metal affinity chromatography. The LC-ESI-MS/MS analysis and a comparison of the amino acid composition demonstrated that the final product was α2-M. The final protein, with a purity of approximately 95% and a yield of nearly 45%, was obtained from Cohn Fraction IV regardless of plasma haptoglobin type, although all but type 1-1 have previously been considered unfavorable for α2-M preparation. The effects of temperature, pH, and methylamine on α2-M activity were evaluated to avoid activity loss during preparation and preservation. The results suggested that α2-M activity could be readily inactivated at temperatures above 50°C, at pH levels above 9.0 or below 4.0, or in the presence of methylamine. Cohn Fraction IV is usually discarded as a biological waste product in the human serum albumin production process; because the simple process developed in this study is relatively inexpensive, the preparation of α2-M from Cohn Fraction IV may better utilize human plasma, a valuable resource.

Existence of porcine endogenous retrovirus (PERV) hinders pigs to be used in clinical xenotransplantation to alleviate the shortage of human transplants. Chinese miniature pigs are potential organ donors for xenotransplantation in China. However, so far, an adequate level of information on the molecular characteristics of PERV from Chinese miniature pigs has not been available. We described here the cloning and characterization of full-length proviral DNA of PERV from Chinese Wuzhishan miniature pigs inbred (WZSP). Full-length nucleotide sequences of PERV-WZSP and other PERVs were aligned and phylogenetic tree was constructed from deduced amino-acid sequences of env. The results demonstrated that the full-length proviral DNA of PERV-WZSP belongs to gammaretrovirus and shares high similarity with other PERVs. Sequence analysis also suggested that different patterns of LTR existed in the same porcine germ line and partial PERV-C sequence may recombine with PERV-A sequence in LTR.

We conducted a large-scale survey on the existence and expression status of porcine endogenous retrovirus (PERV) in seven breeds of Chinese miniature pigs. Genotyping of PERV was examined by PCR using type-specific primers according to the env genotyping method. The presence and expression status of viral gag, pol and env genes were further analyzed in Wuzhishan pigs (WZSP) and Bama minipigs (BMP). The results showed that PERV existed in all 348 genomic DNA samples. The genotype distribution was subtype A-74.43%, subtype B-95.40% and subtype C-30.46%. No expression of subtype C was found in WZSP and BMP. This research obtained an adequate level of information on the molecular epidemiology of PERV in China. The results indicated that it is possible to monitor pig herds for individuals with the lowest PERV prevalence, especially lacking PERV-C.

Background Human parvovirus B19 (B19V) is a frequent contaminant of blood and plasma-derived medicinal products. To ensure the quality and safety of plasma-derived products, European regulations, Plasma Protein Therapeutics Association (PPTA) standard and FDA guidelines require testing of manufacturing plasma for parvovirus B19 DNA to limit the load of this virus. In China, however, there have been no related documentation and technical guiding principles for monitoring B19V, moreover, an adequate level of information on the prevalence of B19V in Chinese plasma donations is not available. Findings By using an in-house quantitative polymerase chain reaction (qPCR) assay adapted for all three genotypes of B19V, 235 source plasma pools from three regional different Chinese manufacturers of blood products were screened and quantified. Results showed that 71.91 % (169/235) of plasma pools were contaminated by B19V, with the concentrations of 5.18 × 102–1.05 × 109 IU/mL. Approximately 31.95 % of the DNA-positive plasma pools were only moderately contaminated (104 IU/mL. Conclusions The high level of B19V in plasma pools could present a great risk in plasma derivatives. Therefore, the implementation of B19V NAT (Nucleic Acid Testing) assays capable of detecting all B19V genotypes and discard donations with high titer B19V DNA for Chinese blood products manufacturers seems to be necessary.

A gold nanoparticle (GNP) probe-based assay (GNPA) modified from the bio-barcode assay (BCA) was developed for ultrasensitive detection of ricin, a potential biothreat agent. In the GNPA, a chain of ricin was captured by a GNP probe coated with polyclonal antibodies and single-stranded signal DNA. A magnetic microparticle (MMP) probe coated with ricin A chain monoclonal antibody was then added to form an immuno-complex. After being magnetically separated, the immuno-complex containing the single-stranded signal DNA was characterized by PCR and real-time PCR. A detection limit of 10(-2) fg/ml was determined for the ricin A chain; this is eight orders of magnitude more sensitive than that achieved with an ELISA and two orders more sensitive than that obtained with the BCA. The coefficients of variation (CV) of the intra- and inter-assay values ranged from 3.82-6.46%. The results here show that this novel assay is an ultrasensitive method for detection of ricin proteins and may be suitable for the ultrasensitive detection of other proteins.

Objective To provide an anatomic basis of the occipital transtentorial keyhole approach (OTKA), then explore its feasibility and surgical indication. Methods Eight cadaveric heads were prepared for this anatomic study. A longitudinal linear 4-cm skin incision that begun at the upper margin of the transverse sinus, 1.5 cm away from the superior sagittal sinus. This was designed for the OTKA. The keyhole craniotomy and conventional craniotomy were performed sequentially for observation and measurement. Results The interhemispheric corridor and the supratentorial corridor can be used in the OTKA. The surgical field extended superior to the splenium, inferior to the superior medullary velum, ipsilateral to the middle and posterior parts of the medial and inferior temporal lobe, contralateral to the pulvinar, and anterior to the massa intermedia in the third ventricle. The exposure area of the OTKA was 72.05 ± 6.26 mm 2 and 182.97 ± 14.65 mm 2 before and after the tentorial incision, respectively. The exposure area of the conventional craniotomy was 187.28 ± 20.16 mm 2 , which had no significant difference to the OTKA. The working angles of the five target points were all smaller for the OTKA than for the conventional approach. The depth of the posterior third ventricle that could be observed was 14.70 ± 2.54 mm with the OTKA. Conclusions Compared with the conventional approach, the OTKA is a more minimally invasive surgical procedure for treatment of the lesions in the pineal region and the middle and posterior parts of the medial and inferior temporal lobe. However, the working angles are relatively narrow.

Gliomas are the most common primary tumors of the central nervous system. Rapid proliferation and diffuse brain invasion of these tumors are likely to determine the unfavorable prognosis. Recent studies have shown that ligand activation of peroxisome proliferator-activated receptor γ (PPARγ) can induce differentiation and inhibit proliferation of several cancer cells. In this study, we identified pioglitazone, one PPARγ ligand in particular, suppressed human glioma cells proliferation, migration, and induced glioma cells apoptosis. Concomitantly, expression level of β-catenin protein, a key molecule in carcinogenesis, was decreased in glioma cells treated with pioglitazone. Noteworthy, knockdown of β-catenin expression using siRNA technology mimicked the anti-neoplastic potency of pioglitazone. These results indicate that β-catenin is one of the mediators for pioglitazone to suppress glioma cells growth and invasion. Due to its capacity to counteract β-catenin and glioma cell proliferation and migration, pioglitazone represents a promising drug for adjuvant therapy of glioma and other highly migratory tumor entities.

BACKGROUND: A recent study provided evidence that farmed rabbits in China harbor a novel hepatitis E virus (HEV) genotype. Although the rabbit HEV isolate had 77-79% nucleotide identity to the mammalian HEV genotypes 1 to 4, their genomic organization is very similar. Since rabbits are used widely experimentally, including as models of infection, we investigated whether they constitute an appropriate animal model for human HEV infection. METHODS: Forty-two SPF rabbits were divided randomly into eleven groups and inoculated with six different isolates of rabbit HEV, two different doses of a second-passage rabbit HEV, and with genotype 1 and 4 HEV. Sera and feces were collected weekly after inoculation. HEV antigen, RNA, antibody and alanine aminotransferase in sera and HEV RNA in feces were detected. The liver samples were collected during necropsy subject to histopathological examination. FINDINGS: Rabbits inoculated with rabbit HEV became infected with HEV, with viremia, fecal virus shedding and high serum levels of viral antigens, and developed hepatitis, with elevation of the liver enzyme, ALT. The severity of disease corresponded to the infectious dose (genome equivalents), with the most severe hepatic disease caused by strain GDC54-18. However, only two of nine rabbits infected with HEV genotype 4, and none infected with genotype 1, developed hepatitis although six of nine rabbits inoculated with the genotype 1 HEV and in all rabbits inoculated with the genotype 4 HEV seroconverted to be positive for anti-HEV IgG antibody by 14 weeks post-inoculation. CONCLUSIONS: These data indicate that rabbits are an appropriate model for rabbit HEV infection but are not likely to be useful for the study of human HEV. The rabbit HEV infection of rabbits may provide an appropriate parallel animal model to study HEV pathogenesis.