Sarcomatoid hepatocellular carcinoma: A case report and review of the literature.

RATIONALE: Sarcomatoid hepatocellular carcinoma (SHC) is a rare malignant tumor composed of both carcinoma and sarcoma components. It has atypical clinical symptoms and a high degree of malignancy, with rapid progression and a poor prognosis. PATIENT CONCERNS: A 63-year-old female patient was admitted to our hospital with a chief complaint of fatigue present for more than 1 month and fever for 10 days. DIAGNOSES: This patient underwent an upper abdominal MRI plain scan and enhanced scan showed a solid tumor in the right lobe of the liver, with a size of approximately 4.7 cm × 4.0 cm × 6.5 cm, present as low signal on T1WI, slightly high signal on T2WI, and heterogeneous high signal on DWI. Multi-phase dynamic contrast-enhanced MR scan showed significant enhancement in the arterial phase and low enhancement in the portal and delayed phases. The pathology showed the tumor cells to be positive for cytokeratin (CK), Vimentin, EMA, CD34, cyclinD1, negative for CK8, CK19, CK20, SMA, Desmin, S-100, CD117, Dog-1, Hepar-1, SOX-10 and ALK, and Ki-67 approximately 50%, which confirmed the diagnosis of SHC. INTERVENTIONS: Laparoscopic right posterior lobe of liver resection was conducted, and the postoperative pathology revealed the presence of SHC. OUTCOMES: The patient was discharged 9 days after the surgery without any complications. There has been no evidence of recurrence at the 1 month, however bilateral pleural metastases appeared during the follow-up 3 months after surgery. LESSONS: SHC is a rare and aggressive liver cancer. So far, there is still a lack of effective therapeutic strategy, and the prognosis was dismal even though patients received radical surgical resection.

Ligamentum teres hepatis as a graft for portal and/or superior mesenteric vein reconstruction: From bench to bedside.

BACKGROUND: Pancreaticoduodenectomy combined with portal vein (PV) and/or superior mesenteric vein (SMV) resection in patients with pancreaticobiliary malignancy has become a common surgical procedure. There are various grafts currently used for PV and/or SMV reconstruction, but each of these grafts have certain limitations. Therefore, it is necessary to explore novel grafts that have an extensive resource pool, are low cost with good clinical application, and are without immune response rejection or additional damage to patients. AIM: To observe the anatomical and histological characteristics of the ligamentum teres hepatis (LTH) and evaluate PV/SMV reconstruction using an autologous LTH graft in pancreaticobiliary malignancy patients. METHODS: In 107 patients, the post-dilated length and diameter in resected LTH specimens were measured. The general structure of the LTH specimens was observed by hematoxylin and eosin (HE) staining. Collagen fibers (CFs), elastic fibers (EFs), and smooth muscle (SM) were visualized by Verhoeff-Van Gieson staining, and the expression of CD34, factor VIII-related antigen (FVIIIAg), endothelial nitric oxide synthase (eNOS), and tissue type plasminogen activator (t-PA) were detected using immunohistochemistry in LTH and PV (control) endothelial cells. PV and/or SMV reconstruction using the autologous LTH was conducted in 26 patients with pancreaticobiliary malignancies, and the outcomes were retrospectively analyzed. RESULTS: The post-dilated length of LTH was 9.67 ± 1.43 cm, and the diameter at a pressure of 30 cm H2O was 12.82 ± 1.32 mm at the cranial end and 7.06 ± 1.88 mm at the caudal end. Residual cavities with smooth tunica intima covered by endothelial cells were found in HE-stained LTH specimens. The relative amounts of EFs, CFs and SM in the LTH were similar to those in the PV [EF (%): 11.23 ± 3.40 vs 11.57 ± 2.80, P = 0.62; CF (%): 33.51 ± 7.71 vs 32.11 ± 4.82, P = 0.33; SM (%): 15.61 ± 5.26 vs 16.74 ± 4.83, P = 0.32]. CD34, FVIIIAg, eNOS, and t-PA were expressed in both LTH and PV endothelial cells. The PV and/or SMV reconstructions were successfully completed in all patients. The overall morbidity and mortality rates were 38.46% and 7.69%, respectively. There were no graft-related complications. The postoperative vein stenosis rates at 2 wk, 1 mo, 3 mo and 1 year were 7.69%, 11.54%, 15.38% and 19.23%, respectively. In all 5 patients affected, the degree of vascular stenosis was less than half of the reconstructed vein lumen diameter (mild stenosis), and the vessels remained patent. CONCLUSION: The anatomical and histological characteristics of LTH were similar to the PV and SMV. As such, the LTH can be used as an autologous graft for PV and/or SMV reconstruction in pancreaticobiliary malignancy patients who require PV and/or SMV resection.

Counterclockwise modular laparoscopic anatomical mesohepatectomy using combined glissonean pedicle (Takasaki approach) and hepatic vein-guided approaches.

Background: Although laparoscopic anatomical hepatectomy (LAH) is widely adopted today, laparoscopic anatomic mesohepatectomy (LAMH) for patients with hepatocellular carcinoma (HCC) remains technically challenging. Methods: In this study, 6 patients suffering from solitary liver tumors located in the middle lobe of the liver underwent counterclockwise modular LAMH using combined Glissonean pedicle (Takasaki approach) and hepatic vein-guided approaches. In this process, the Glissonean pedicle approach (Takasaki approach) was first used to transect the liver pedicles of segment right anterior (G58) and segment 4 (G4). Second, the hepatic vein-guided approach was performed along the umbilical fissure vein (UFV) to sever the liver parenchyma from the caudal to cranial direction, and the middle hepatic vein (MHV) and anterior fissure vein (AFV) were then disconnected at the root. Last, the hepatic vein-guided approach was once more performed along the ventral side of the right hepatic vein (RHV) to transect the liver parenchyma from the cranial to anterior direction, and the middle lobe of the liver, including the tumor, was removed completely. The entire process was applied in a counterclockwise fashion, and the exposure or transection sequence was G58, and G4, followed by UFV, MHV, AFV, and finally, the liver parenchyma along the ventral side of RHV. Results: The counterclockwise modular LAMH using combined Glissonean pedicle (Takasaki approach) and hepatic vein-guided approaches was feasible in all 6 cases. The median duration of the operation was 275 ± 35.07 min, and the mean estimated blood loss was 283.33 ml. All of the 6 patients recovered smoothly. The Clavien-Dindo Grade I-II complications rate was up to 33.33%, mainly characterized by postoperative pain and a small amount of ascites. No Clavien-Dindo Grade III-V complications occurred, and the mean postoperative hospital stay was 6.83 ± 1.47 days. Follow-up results showed that the average disease-free survival (DFS) was 12.17 months, and the 21-months OS rate, DFS rate and tumor recurrent rate were 100%, 83.33% and 16.67% respectively. Conclusions: Counterclockwise modular LAMH using combined Glissonean pedicle (Takasaki approach) and hepatic vein-guided approaches takes the advantages of the two approaches, is a novel protocol for LAMH. It is thought to be technically feasible for patients with a centrally located solitary HCC. The oncologic feasibility of this technique needs to be investigated based on long-term follow-up. A multicenter, large-scale, more careful study is necessary.

PGC-1α Protects against Hepatic Ischemia Reperfusion Injury by Activating PPARα and PPARγ and Regulating ROS Production.

Peroxisome proliferator-activated receptors (PPARs) α and γ have been shown to be protective in hepatic ischemia/reperfusion (I/R) injury. However, the precise role of PPARγ coactivator-1α (PGC-1α), which can coactivate both of these receptors, in hepatic I/R injury, remains largely unknown. This study was designed to test our hypothesis that PGC-1α is protective during hepatic I/R injury in vitro and in vivo. Our results show that endogenous PGC-1α is basally expressed in normal livers and is moderately increased by I/R. Ectopic PGC-1α protects against hepatic I/R and hepatocyte anoxia/reoxygenation (A/R) injuries, whereas knockdown of endogenous PGC-1α aggravates such injuries, as evidenced by assessment of the levels of serum aminotransferases and inflammatory cytokines, necrosis, apoptosis, cell viability, and histological examination. The EMSA assay shows that the activation of PPARα and PPARγ is increased or decreased by the overexpression or knockdown of PGC-1α, respectively, during hepatic I/R and hepatocyte A/R injuries. In addition, the administration of specific antagonists of either PPARα (MK886) or PPARγ (GW9662) can effectively decrease the protective effect of PGC-1α against hepatic I/R and hepatocyte A/R injuries. We also demonstrate an important regulatory role of PGC-1α in reactive oxygen species (ROS) metabolism during hepatic I/R, which is correlated with the induction of ROS-detoxifying enzymes and is also dependent on the activations of PPARα and PPARγ. These data demonstrate that PGC-1α protects against hepatic I/R injury, mainly by regulating the activation of PPARα and PPARγ. Thus, PGC-1α may be a promising therapeutic target for the protection of the liver against I/R injury.

Correlation between Intraoperative Fluid Administration and Outcomes of Pancreatoduodenectomy.

BACKGROUND: Intraoperative fluid (IOF) administration plays an important role during major abdominal surgery although increased fluid intake can adversely influence postoperative outcomes. However, the effect of the IOF rate on the outcomes of pancreatoduodenectomy (PD) is unclear. METHODS: 151 patients, who underwent PD at Binzhou Medical University Hospital between January 2010 and May 2017, were categorized into three groups according to IOF rates (ml/kg/hr): restricted (<10, n = 47), standard (10-15, n = 76), and liberal (>15, n = 28). RESULTS: The overall postoperative morbidity was 56.95%. The incidence of postoperative pancreatic fistula (POPF) was 11.26%. The in-hospital mortality rate was 7.28% with the most common cause being grade C POPF and secondary intra-abdominal infections. The patients in the liberal group had significantly higher incidences of POPF (25%) and respiratory complications (21.43%). The other outcome parameters such as recovery of bowel function, hospital stay, and postoperative daily drainage were similar among the groups. Multivariable analysis confirmed the IOF rate to be most strongly associated with POPF (odds ratio: 5.195, confidence interval: 1.142-23.823, P = 0.023) and respiratory complications (odds ratio: 7.302, confidence interval: 0.676-58.231, P = 0.025). CONCLUSIONS: The IOF rate significantly affects the incidence of POPF and respiratory complications after PD. Careful patient-oriented fluid therapy may help to prevent these complications.

PCV cap proteins fused with calreticulin expressed into polymers in Escherichia coli with high immunogenicity in mice.

BACKGROUND: Porcine circovirus type 2 (PCV2) is the main causative agent of porcine circovirus diseases (PCVDs) which causes huge yearly economic losses in the swine industry. Capsid protein (Cap) is the major structural protein of PCV2 that can induce a protective immune response. Therefore, developing a novel and safe subunit vaccine against PCV2 infection is needed. RESULTS: In this study, the Cap gene was bound to the truncated calreticulin (CRT) (120-250 aa/120-308 aa) at the N/C terminal, and then the CRT-Cap fusion genes were expressed in Escherichia coli (E.coli). The size-exclusion chromatography and dynamic light scattering (DLS) data showed that the purified recombinant CRT-Cap fusion protein (rP5F) existed in the form of polymers. Immunization with rP5F stimulated high levels of PCV2 specific antibody and neutralization antibody in mice, which were almost identical to those induced by the commercial subunit and inactivated vaccines. The lymphocyte proliferation and cytokine secretion were also detected in rP5F immunized mice. According to the results of PCV2-challenge experiment, the virus loads significantly decreased in mice immunized with rP5F. The data obtained in the current study revealed that rP5F had the potential to be a subunit vaccine candidate against PCV2 in the future. CONCLUSIONS: We have successfully expressed Cap-CRT fusion proteins in E.coli and optimized rP5F could form into immunogenic polymers. Mice immunized with rP5F efficiently induced humoral and part of cellular immune responses and decreased the virus content against PCV2-challenge, which suggested that rF5P could be a potential subunit vaccine candidate.

Pancreatoduodenectomy for Periampullary Tumors Presenting with Acute Pancreatitis.

BACKGROUND: Periampullary tumors (PT) may rarely present as acute pancreatitis (AP) or acute recurrent pancreatitis (ARP). Unlike other cases of AP and ARP, these conditions necessitate pancreaticoduodenectomy (PD), and timely diagnosis is crucial. Materials and Methods. A retrospective review of clinical, radiological, surgical, and pathological data was conducted for patients admitted to the Binzhou Medical University Hospital during the period from January 2010 to December 2017, for AP or ARP caused by PT. All patients included in the study group had undergone PD. The perioperative data for these patients was compared with data for patients with PT but without AP or ARP who underwent PD during the same period (control group). RESULTS: During the study period, 412 patients with AP or ARP were treated; among this group, 15 patients had PT. Compared with controls, patients in the study group were younger in age and had a longer course of disease, more frequent hospitalizations, and more severe derangements in laboratory data (P < 0.05). Operative time and intraoperative blood loss were significantly higher in the study group, but the incidence of postoperative outcomes such as pancreatic/biliary fistula, abdominal infection, postoperative hospital stay, and mortality were similar between groups (P < 0.05). Operative time and intraoperative blood loss were significantly higher in the study group, but the incidence of postoperative outcomes such as pancreatic/biliary fistula, abdominal infection, postoperative hospital stay, and mortality were similar between groups (. CONCLUSIONS: Neither AP nor ARP has any adverse impact on the outcomes of PD. However, in the treatment of younger patients suffering from AP or ARP, unexplained pancreatic duct dilation and weight loss should raise the suspicion of PT. EUS and EUS-FNA may be helpful in making the diagnosis.

Ethanol extract of Acanthopanax senticosus (Rupr. & Maxim.) Harms induces liver cancer cell apoptosis through inhibiting NF-κB.

BACKGROUND: Acanthopanax senticosus (Rupr. & Maxim.) Harms, a traditional Chinese medicine, has been used to treat various diseases, including ischemic, heart diseases, hepatocellular carcinoma (HCC), hypertension and neurasthenia. The purpose of this study was to investigate the anticancer activity of A. senticosus (ASE). MATERIALS: MTT assay, clonogenicity, reverse transcription-polymerase chain reaction (RT-PCR), flow cytometry and Western blot were employed to evaluate the viability and invasion of liver cancer cells. In addition, luciferase assay was used to delineate the inhibitory activity of ethanol extract against NF-κB. RESULTS: Our results showed that the ethanol extract of ASE could decrease the viability of cancer cells. In addition, the ethanol extract could decrease the protein levels of Matrix metalloproteinase-2 (MMP-2), MMP-9, t-protein kinase B (Akt) and p-Akt, but increase those of E-cadherin. Nuclear factor kappa beta (NF-κB)-Luciferase assay showed the ethanol extract could effectively inhibit the activity of NF-κB. Furthermore, fourteen compounds including seven active compounds were isolated and activities against NF-κB were investigated. CONCLUSIONS: This study confirmed that the ASE could be as an alternative or complementary therapy to treat liver cancer.

Retraction: Ethanol extract of Acanthopanax senticosus (Rupr. & Maxim.) Harms induces liver cancer cell apoptosis through inhibiting NF-κB.

[This retracts the article DOI: 10.21037/tcr.2020.01.02.].

Spleen-preserving distal pancreatectomy and lymphadenectomy for glucagonoma syndrome: A case report.

RATIONALE: Glucagonoma is a rare type of functional pancreatic neuroendocrine tumor that is characterized by distinctive clinical manifestations; among these, necrolytic migratory erythema represents the hallmark clinical sign of glucagonoma syndrome and is usually presented as the initial complaint of patients. PATIENT CONCERNS: A 30-year-old male patient was admitted to our hospital with a complaint of diffuse erythematous ulcerating skin rash for more than 10 months. He also complained of hyperglycemia and a weight loss of 15 kg in those months. DIAGNOSIS: This patient underwent a contrast-enhanced computed tomography scan which showed a pancreatic body mass measuring approximately 6 cm with low density accompanied by partial calcification in plain scanning images and uneven enhancement in strengthening periods. In addition, laboratory tests indicated elevated fasting blood glucagon (1109 pg/mL, normal range: 50-150 pg/mL) levels. Glucagonoma syndrome was ultimately diagnosed in clinical. INTERVENTION: Spleen-preserving distal pancreatectomy was conducted and postoperative pathology revealed the presence of glucagonoma. OUTCOMES: The patient recovered uneventfully with the glucagonoma syndrome disappeared soon after surgery, and the postoperative plasma glucagon decreased to a normal level. Follow-up showed no recurrence for 5 years since the surgery. LESSONS: The treatment of glucagonoma should be directed according to the stage at which the disease is diagnosed. Surgery is currently the only method available to cure the tumor, although medications are given to patients who present with advanced glucagonoma and who are not candidates for operation. Multidisciplinary therapy and multimodality treatment are advised, although these have been systematically evaluated to a lesser degree.

MiRNA-124 inhibits the proliferation, migration and invasion of cancer cell in hepatocellular carcinoma by downregulating lncRNA-UCA1.

Purpose: It has been reported that miRNA-124 inhibits hepatocellular carcinoma (HCC) progression, while lncRNA-UCA1 promotes HCC. The aim of this study is to find whether miRNA-124, as a tumor suppressor in HCC can inhibit lncRNA-UCA1 in HCC cell. Methods: Tumor tissues and adjacent healthy tissues were obtained from 66 patients diagnosed with HCC in Binzhou Medical University Hospital from January 2011 to January 2013. Cell proliferation assay, in vitro cell migration and invasion assay were applied for the research. Results: In the present study we found that miRNA-124 was downregulated, while lncRNA-UCA1 was upregulated in tumor tissues comparing to adjacent healthy tissues of HCC patients. Expression of miRNA-124 and lncRNA-UCA1 in tumor tissues were not affected by HBV or HCV infection. Analysis of followed-up data revealed that low miRNA-124 level and high lncRNA-UCA1 level were closely correlated with poor survival. Overexpression of miRNA-124 led to inhibited lncRNA-UCA1 expression in cells of HCC cell lines, while overexpression of lncRNA-UCA1 failed to significantly affect miRNA-124 expression. Expression levels of miRNA-124 and lncRNA-UCA1 were inversely and significantly correlated in tumor tissues but not in adjacent healthy tissues. Overexpression of miRNA-124 led to inhibited, while overexpression of lncRNA-UCA1 led to increased proliferation, migration and invasion rates of HCC cell lines. In addition, lncRNA-UCA1 overexpression attenuated the inhibitory effects of miRNA-124 overexpression on cancer cell proliferation, migration and invasion. Conclusion: Therefore, miRNA-124 may inhibit the proliferation, migration and invasion of cancer cell in hepatocellular carcinoma by downregulating lncRNA-UCA1.

A giant mesenteric fibromatosis involving the muscular layer of the colon wall: A case report.

RATIONALE: Mesenteric fibromatosis (MF) is a rare tumor whose biological behavior is intermediate between benign fibrous neoplasms and fibrosarcomas, and the characteristic of these tumors are local aggressive lesions which is prone to local recurrence but non-metastasizing. The common symptom is abdominal distention or painless mass. We report a case of giant MF in abdominal cavity with abdominal distention as the main symptom. PATIENT CONCERNS: A 26-year-old male presented with 2-month history of abdominal distention, lack of appetite, and symptoms grew progressively more debilitating with time. DIAGNOSES: This patient underwent a contrast-enhanced computed tomography scan which showed a giant (37 × 25 × 13 cm), inhomogeneous enhancing, well-defined, and soft tissue density mass in abdominal cavity, possibly arising in mesocolon, which suggested a high possibility of MF. The postoperative pathology showed that the tumor cells to be positive for ß-catenin, vimentin, negative for CD34, CD117, DOG-1, S-100, Desmin, which confirmed the diagnosis of MF. INTERVENTIONS: Exploratory laparotomy was performed, which revealed a large mass involving the transverse colon wall, the root of mesocolon, and encasing the middle colic vessels and the 1st branch of jejunal arteries. The complete surgical resection was performed and the mass weighted 10 kilograms (kg). OUTCOMES: The patient recovered uneventfully and was discharged 9 days after surgery. Three-month, 6-month, 12-month and 18-month on follow-up after surgery, showed no evidence of recurrence. LESSONS: The MF is a very rare tumor, especially a giant tumor (10 kg) involving the muscular layer of colon wall. In addition, treatment of giant MF still remains a challenge. We consider that surgical resection with negative margins is the goal but not at the expense of damaging the function of vital organs. Specific measures should be considered based on the individual patient in order to relieve symptoms and improve quality of life.

MicroRNA-187 inhibits tumor growth and metastasis via targeting of IGF-1R in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the primary and most frequently occurring type of malignant liver cancer, accounting for 70-85% of total liver cancer cases worldwide. It has previously been demonstrated that the aberrant expression of microRNAs (miR) contributes to carcinogenesis and progression of various human malignancies, including HCC. However, mechanisms underlying the differential expression and specific roles of miR­187 in HCC remain to be elucidated, particularly regarding how the modulation of malignant phenotypes in HCC cells occurs. The present study demonstrated that miR­187 was significantly downregulated in HCC tissues and cell lines. Restoration of miR­187 expression inhibited cell proliferation, migration and invasion in HCC. Furthermore, insulin­like growth factor 1 receptor (IGF­1R) was demonstrated to act as a direct target gene of miR­187 in HCC. IGF­1R knockdown mimicked the effects of miR­187 overexpression in HCC, resulting in a significant inhibition of cell proliferation, migration and invasion. The results of the present study demonstrated that miR­187 acted as a tumor suppressor in HCC progression via direct targeting of IGF­1R. miR­187 may therefore exhibit the potential to act as a novel and therapeutic target for HCC treatment in the future.

Molecular epidemiology of porcine reproductive and respiratory syndrome virus in Central China since 2014: The prevalence of NADC30-like PRRSVs.

Porcine reproductive and respiratory syndrome (PRRS), characterized by respiratory disorders in piglets and reproductive failure in sows, is still the great threat of swine industry. Recently, Emergence of the novel NADC30-like PRRS viruses (PRRSVs) has caused widespread outbreaks of PRRS. To investigate the epidemic characteristics of PRRSVs in Central China since 2014, 6372 clinical serum samples were tested by ELISA, 250 tissue samples were tested by RT-PCR, and among these, 30 ORF5 and 17 Nsp2 genes sequences were analyzed. Phylogenetic tree based on ORF5 revealed that, 17 isolates were clustered into subgroup 1, represented by the NADC30. And for the Nsp2, The strains which had a discontinuous 131-amino-acid deletion in Nsp2, called NADC30-like strains, were clustered into subgroup 2. Our data suggested that the NADC30-like PRRSV strains spread quickly and are now circulating and prevalent in Central China as well as the classical HP-PRRSV strains. In addition, amino acid variation analysis of GP5 revealed that the amino acid sequences of NADC30-like PRRSV strains underwent rapid evolution and contained extensive amino acid substitutions in important motifs, such as potential neutralization epitope and the N-glycosylation sites. In summary, our data would provide a large amount of detailed information on molecular variation and genetic diversity of PRRSV in central China.

Reduction of hepatic fibrosis by overexpression of von Hippel-Lindau protein in experimental models of chronic liver disease.

Hypoxia-inducible factor (HIF)-1α and HIF-2α play an important role in liver fibrosis. von Hippel-Lindau protein (VHL), a key mediator of HIF-α, regulates fibrosis in an organ- and cell-specific way. In this study, human liver samples were collected from hepatitis C-, alcoholic-, and cholestatic-associated fibrotic and healthy individuals. Two mouse models of liver fibrosis were established: bile duct ligation and carbon tetrachloride injection. We constructed adenovirus vectors to overexpress VHL, normoxia-active HIF-α, and lentiviral vectors to silence HIF-α. The results showed that liver sections from fibrosis patients had a lower level of VHL and higher levels of HIF-1α and HIF-2α compared with healthy sections, a finding which was confirmed in mice. Overexpression of VHL attenuated liver fibrosis, downregulated fibrogenic genes, and inhibited liver inflammation, apoptosis, and angiogenesis. Overexpression of VHL was more successful at inhibiting fibrosis compared with silencing HIF-1α plus HIF-2α. Normoxia-active HIF-1α or HIF-2α prevented the inhibitory effect of VHL on liver fibrosis, indicating that attenuating fibrosis via VHL is HIF-1α- and HIF-2α-dependent to some extent. In addition, overexpression of VHL inhibited mouse hepatic stellate cells activation and proliferation and promoted apoptosis. Taken together, VHL may be considered a new target to inhibit liver fibrosis.

Porcine parvovirus capsid protein expressed in Escherichia coli self-assembles into virus-like particles with high immunogenicity in mice and guinea pigs.

Porcine parvovirus (PPV) is a causative agent of reproductive failure in pregnant sows. Classical inactivated vaccine is extensively used to control PPV infection, but problems concerning safety, such as incomplete inactivation may occur. In this study, a novel subunit vaccine against PPV based on virus-like particles (VLPs) formed from the complete PPV VP2 protein expressed in a prokaryotic system with co-expressed chaperones is reported. The VLPs have a similar size, shape, and hemagglutination property to the PPV. Immunization with these VLPs stimulated the neutralization antibody and hemagglutination inhibition (HI) antibody responses in mice and guinea pigs. The lymphocyte proliferation response and cytokine secretion was also induced in immunized guinea pigs comparable to those immunized with PPV inactivated vaccine. In addition, immunization with VLPs also significantly reduced the PPV content in the spleen of guinea pigs 14 days after the challenge with intact virus. These studies suggest that PPV VLPs created as described here could be a potential candidate for vaccine development.

Detection and phylogenetic analysis of porcine epidemic diarrhea virus in central China based on the ORF3 gene and the S1 gene.

BACKGROUND: Porcine epidemic diarrhea (PED) has increased in severity in China since 2010. To investigate further the infectivity, genetic diversity and molecular epidemiology of its causative agent, the porcine epidemic diarrhea virus (PEDV), we assessed 129 clinical samples, which were the intestinal tissue of piglets with severe diarrhea, from 17 cities in central China. Both the spike (S) glycoprotein (S1, 1-789 amino acids (aa)) and the full-length ORF3 gene of 21 representative field strains from 21 farms in 11 cities were sequenced and analysed. METHODS: PEDV was detected by reverse transcription-polymerase chain reaction (RT-PCR), and S1 and ORF3 sequences were processed by the Clustal W method via DNAMAN 8 software, and phylogenetic trees were constructed by the neighbor-joining method using MEGA 6 software. RESULTS: The prevalence of PEDV was 92.25% and was detected in 119 of 129 samples, with 94.03% (63 of 67) of pig farms harbouring the disease. According to the phylogenetic analysis of the S1 genes, our isolates all fell into group G2 (variants) and showed a close relationship to isolates from Chinese (HN1303, CH/ZMDZY/11 and AJ1102), Korean (AD01), American (MN, IA1, IA2 and 13-019349) sources, and these isolates differed genetically from other Chinese (LZC, CH/HNZZ/2011 and SD-M) and Korean (SM98) strains as well Japanese (83-P5 and MK) strains. In addition, our isolates differed from attenuated vaccine strains, CV777 (used in China) and DR13 (used in Korea). According to our derived amino acid sequence analysis, we detected one novel variant PEDV, viz: CH/HNLY, with 4-aa insertion/deletion (RSSS/T) at position 375 and 1-aa (D) deletion at position 430 compared to the CV777 attenuated strain. These mutations were located on the receptor binding domain. Our ORF3 gene analyses showed that the prevalent PEDV isolates were variants, and the isolated strains differed genetically from the vaccine strains. CONCLUSIONS: These findings illustrated the existence of genetic diversity among geographically distinct PEDV strains, and our study has provided an impetus to conduct further research on the PEDV receptor binding protein and on the new and efficacious vaccines design.

Human papillomavirus L1 protein expressed in Escherichia coli self-assembles into virus-like particles that are highly immunogenic.

HPV vaccines based on L1 virus-like particles (VLPs) provided a high degree of protection against HPVs infection. In this study, the codon optimized HPV16 L1 gene were sub-cloned into five procaryotic expression vectors (pET-28a, pET-32a, pGEX-4T-2, pE-sumo and pHSIE), and fused with different protein tags. No recombinant proteins were expressed in pET-28a-L1 and pHSIE-L1, and the proteins expressed by pET-32a-L1 plasmid with TRX-tag were in the form of inclusion body. Only SUMO-tagged and GST-tagged L1 proteins expressed by pE-Sumo-L1 or pGEX-4T-L1 were soluble. The yield of SUMO-L1 protein reached 260mg/L fermentation medium in shake flask. After SUMO tags were eliminated, a 90% purity of L1 proteins was generated by ion-exchange and Ni-NTA affinity chromatography. The purified HPV16 L1 protein self-assembled into virus-like particles (VLPs) and showed a haemagglutination activity. High titers specific and neutralizing antibodies were detected in HPV 16 L1VLPs vaccinated mice. Cytokines such as IFN-γ and IL-2 showed significant higher in VLPs vaccinated mice compared with negative control (p<0.05, p=0.055). Thus, the expression of recombinant HPV16 L1 VLPs in Escherichia coli was feasible, which could potentially be used for a VLP-based HPV vaccine.

High level soluble expression and one-step purification of IBDV VP2 protein in Escherichia coli.

OBJECTIVES: To improve the expression of soluble IBDV VP2 protein by using different tagged vectors in Escherichia coli. RESULTS: Fusion tags, Grifin, MBP, SUMO, thioredoxin, γ-crystallin, ArsC and PpiB, enhanced the expression and solubility of VP2 protein. The fusion proteins were purified by Ni-NTA chromatography, MBP-VP2 showed the highest purity about 90 %. After removing the MBP tag, VP2 self-assembled into virus-like particles, ~25 nm diam. Results from AGP suggested the recombinant IBDV VP2 protein identified by reference serum like IBDV. CONCLUSION: All the seven tags enhanced the expression and solubility of IBDV VP2 protein. The recombinant protein self-assembly into virus like particles and possess antigenicity as reference IBDV.

Sodium orthovanadate inhibits growth of human hepatocellular carcinoma cells in vitro and in an orthotopic model in vivo.

The transition metal vanadium is widely distributed in the environment and exhibits various biological and physiological effects in the human body. As a well known vanadium compound, sodium orthovanadate (SOV) has shown promising antineoplastic activity in several human cancers. However, the effects of SOV on liver cancer are still unknown. In this study, for the first time, we showed that SOV could effectively suppress proliferation, induce G2/M cell cycle arrest and apoptosis, and diminish the mitochondrial membrane potential (MMP) of HCC cells in vitro. In addition, our in vitro results were recapitulated in vivo, showing that SOV exhibited a dose-dependent inhibition of growth of human HCC in an orthotopic model, evidenced by the reduction in tumor size, proliferation index and microvessel density, and increase in cell apoptosis. Most important, we found that SOV could inhibit autophagy in HCC cells in vitro and in vivo, which plays a prodeath role. Thus, our findings suggest that SOV could effectively suppress the growth of human HCC through the regulations of proliferation, cell cycle, apoptosis and autophagy, and thus may act as a potential therapeutic agent in HCC treatment.