SIRT3 Promotes the Development of Esophageal Squamous Cell Carcinoma by Regulating Hexokinase 2 through the AKT Signaling Pathway.

In the present study, we explored whether sirtuin-3 (SIRT3) regulates the proliferation and migration of esophageal squamous cell carcinoma (ESCC) and investigated the mechanisms underlying the oncogene role of SIRT3. siRNA was used to transfect Eca109 cells and downregulate SIRT3. The proliferation and migration of Eca109 cells were examined by the CCK-8 assay, colony formation assay, Transwell assay, and scratch test. Quantitative real-time PCR and Western blotting were used to detect SIRT3, hexokinase 2, AKT, and p-AKT in Eca109 cells. Functional assays showed that downregulation of SIRT3 could inhibit the proliferation and migration of ESCC cells. Reduced SIRT3 expression downregulated hexokinase 2 expression and inhibited AKT activation in ESCC. These results indicated that SIRT3 promote ESCC development and progression by regulating hexokinase 2 through the AKT signaling pathway. SIRT3 promote ESCC proliferation and migration by regulating HK-2 through the AKT signaling pathway.

Blood service in the Tibetan regions of Garzê and Aba, China: a longitudinal survey.

BACKGROUND: Garzê and Aba form the second largest Tibetan-inhabited area of China. Blood services have never been reported for this region before. OBJECTIVE: To assess the current situation and analyse whether a safe and adequate blood supply has been developed in both Garzê and Aba. METHODS: We conducted a longitudinal survey covering the period 2011-2016. The subjects of interest were recruited from non-remunerated voluntary donation, blood testing, clinical transfusion practices and infrastructure of local blood service systems. RESULTS: The donation rate and blood collection volume were below the average levels of both the Sichuan Province and mainland China. Component therapy was widely used, but inappropriate usage of whole blood existed. A lack of national specific standards for people on the plateaus led to local blood transfusions being conducted without full clinical assessment. Endemic and frequently occurring disease, such as hydatid disease and gastrointestinal disease, were inevitable risks for blood utilisation and safety. The potential influence of religious belief and traditions, like 'male-leaving marriages', of Tibetans on donor recruitment and blood safety requires further research. CONCLUSIONS: A relatively safe and complete blood service system has been developed in this region. However, there is still an urgent need for comprehensive and effective support from the government in terms of policies and finance. As an epidemic area of hydatid disease and sexually transmitted disease, this region needs to emphasise public health measures, such as blood safety and inappropriate usage of blood products.

Application of vacuum-assisted closure in seawater-immersed wound treatment under different negative pressures.

The therapeutic effect of vacuum-assisted closure (VAC) has been confirmed in many types of complex wounds, but there are few relevant reports regarding seawater-immersed wounds. The aim of this study was to determine the effect of VAC on seawater-immersed wound healing under different negative pressures and explore the optimal negative pressure value. Four purebred miniature pigs were used as the experimental animal models. Four acute, symmetrical wounds were made on each side of the spine and designated as the experimental group (wounds with 2 h of seawater immersion) and the control group (wounds without seawater immersion). Wounds were divided into a conventional dressing group and 3 further groups with different VAC therapies (negative pressure at either 120, 180, or 240 mmHg). The extent of wound healing, and speed of granulation growth and re-epithelialization were measured. Bacterial flora distribution in the wounds was observed, and fibronectin levels in the exudate of the wounds were tested. Results showed that seawater immersion aggravated wound injury and that VAC therapy with 180 mmHg negative pressure induced the fastest epidermis migration, obvious edema elimination, significant capillary proliferation, and the highest level of fibronectin, and that in wounds, the proportion of Gram-negative bacteria tended to decrease and that of Gram-positive bacteria tended to increase. Our results show that VAC promotes seawater-immersed wound healing and that 180 mmHg negative pressure may be optimal for wound healing.

Clinical analysis of osteosarcoma patients treated with high-dose methotrexate-free neoadjuvant chemotherapy.

OBJECTIVE: High-dose methotrexate (hdmtx) is a common therapeutic agent in the treatment of osteosarcoma. However, hdmtx is highly toxic and requires complex pharmacokinetic monitoring and leucovorin rescue. Thus, alternative therapeutic strategies are necessary. Here, we analyzed the clinical efficacy of a dia regimen (cisplatin-ifosfamide-doxorubicin) to evaluate its potential as an alternative to hdmtx-based therapy. METHODS: Patients received 12 cycles of chemotherapy administered over 2 years (2 preoperative cycles and 10 postoperative cycles). Cumulative dose was the same in all cycles: cisplatin 120 mg/m(2) on day 1 of week 1, followed by ifosfamide 2.0 g/m(2) days 1-5 of week 2, and doxorubicin 20 mg/m(2) days 1-3 of week 2. RESULTS: Between January 2004 and October 2008, 39 eligible patients (median age: 16 years) were enrolled, with 36 being evaluable for the study. Of those 36 patients, 20 (55.6%) had a good histologic response to preoperative chemotherapy (>90% tumour necrosis). The estimated 5-year rates of event-free survival (efs) and overall survival were 54.8% and 61.5% respectively. CONCLUSIONS: The results of our study suggest that, in osteosarcoma patients, the dia regimen produces an efs rate and survival outcomes comparable to those attained with hdmtx-containing regimens, with fewer adverse reactions. The dia regimen is well tolerated, and we observed a high level of patient compliance. Our results demonstrate that hdmtx-free osteosarcoma treatment regimens can be effective, warranting further investigation.

Successful management of a childhood osteosarcoma with epiphysiolysis and distraction osteogenesis.

In an 11-year-old boy with osteosarcoma in the proximal tibia (type iii), 2 cycles of dia chemotherapy (cisplatin, ifosfamide, doxorubicin) were administered preoperatively while epiphysiolysis was performed. Clinical response was determined to be complete by radiography and histopathology. Marginal excision was then performed with preservation of the proximal tibial epiphysis. Metaphyseal reconstruction was performed using distraction osteogenesis. Six cycles of dia chemotherapy were administered postoperatively. Twenty months later, the patient had developed no complications and experienced full bone healing, with no limb discrepancy. In selected adolescent patients with osteosarcoma, in whom the tumour is in full contact with the epiphysis, epiphyseal preservation by epiphysiolysis and reconstruction by distraction osteogenesis can provide an excellent outcome, resulting in a stable reconstruction that functionally restores the native limb.

Comparison of the diagnostic value of procalcitonin and C-reactive protein after hematopoietic stem cell transplantation: a systematic review and meta-analysis.

OBJECTIVE: To conduct a systematic review and meta-analysis of the performance of the procalcitonin (PCT) diagnostic test for identifying infectious complications after hematopoietic stem cell transplantation (HSCT). METHODS: We searched EMBASE, MEDLINE, the Cochrane database, and reference lists of relevant articles, with no language restrictions, through December 2011. We selected original articles that reported diagnostic performance of PCT alone or compared with other biomarkers for identifying serious infections in HSCT recipients. We quantitatively evaluated test accuracy parameters with the use of forest plots, hierarchical summary receiver operating characteristic curves, and bivariate random effect models. RESULTS: We found 6 qualifying studies (studying 1344 episodes of suspected infection with confirmed infectious episodes) from 3 countries. These 6 studies examined both PCT and C-reactive protein (CRP) test performance. Bivariate pooled sensitivity, specificity, positive likelihood ratios, and negative likelihood ratios were 0.66 (95% confidence interval [CI] 0.60-0.72), 0.72 (95% CI 0.65-0.79), 2.39 (95% CI 1.84- 3.09), and 0.47 (95% CI 0.39-0.57) for PCT, and 0.80 (95% CI 0.54-0.93), 0.73 (95% CI 0.56-0.86), 3.00 (95% CI 1.86-4.84), and 0.27 (95% CI 0.11-0.65) for CRP. In terms of area under the curve (AUC), CRP was superior to PCT in detecting infectious complications, with an AUC of 0.82 for CRP versus an AUC of 0.69 for PCT. CONCLUSION: The pooled accuracy estimates of 6 different studies indicated only a moderate rule-out diagnostic value of both PCT and CRP in discriminating infection from other inflammatory complications following allogeneic HSCT.

Denosumab and giant cell tumour of bone-a review and future management considerations.

Giant cell tumour of bone (gctb) is one type of giant-cell-rich bone lesion characterized by the presence of numerous multinucleated osteoclast-type giant cells. Giant cells are known to express rankl (receptor activator of nuclear factor κB ligand) and are responsible for the aggressive osteolytic nature of the tumour. No available treatment option is definitively effective in curing this disease, especially in surgically unsalvageable cases. In recent years, several studies of denosumab in patients with advanced or unresectable gctb have shown objective changes in tumour composition, reduced bony destruction, and clinical benefit. Denosumab is a fully human monoclonal antibody that targets and binds with high affinity and specificity to rankl. Several large phase iii studies have shown that denosumab is more effective than bisphosphonates in reducing skeletal morbidity arising from a wide range of tumours and that it can delay bone metastasis. The relevant articles are reviewed here. The controversies related to the future use of denosumab in the treatment of gctb are discussed.

[The value of serum heavy/light chain immunoassay to assess therapeutic response in patients with multiple myeloma].

Objective: To assess the value of immunoglobulin heavy/light chain (HLC) immunoassay on therapeutic response in patients with multiple myeloma(MM). Methods: A total of 45 newly diagnosed MM patients were retrospectively enrolled in Peking Union Medical College Hospital from 2013 to 2016, whose 115 serum samples were consecutively collected. HLC was tested to evaluate response and compare with other methods for M protein detection. Results: ①There were 30 males and 15 females in total of whom the monoclonal immunoglobulin was IgG in 27 (IgGκ∶IgGλ 12∶15) and IgA (IgAκ∶IgAλ 9∶9) in 18. The arerage age of the studied population was 59 (range 43-80) . ② In 34 patients with serum sample at diagnosis, 32 (94.1%) had abnormal HLC ratio (rHLC) while 2 patients with IgG had normal rHLC. The percentages of abnormal rHLC was 81.8% (18/22) at partial response、50.0%(9/18) at very good complete response and 16.0%(4/25) at complete response. ③In 25 patients reaching CR, there were 13 with IgG and 12 with IgA. 4 patients equally split of IgG and IgA had abnormal rHLC at complete response. ④By monitoring the rHLC of some patients consecutively, we found that the remission of rHLC was to some extent behind the remission of SPE and IEF, or even rFLC. Conclusion: Immunoglobulin HLC detection is one feasible method for minimal residual disease detection.

Concentration and chemical characteristics of PM2.5 in Beijing, China: 2001-2002.

Weekly PM2.5 samples were simultaneously collected at a semi-residential (Tsinghua University) and a downtown (Chegongzhuang) site in Beijing from August 2001 through September 2002. The ambient mass concentration and chemical composition of PM2.5 were determined. Analyses including elemental composition, water-soluble ions, and organic and elemental carbon were performed. The annual average concentrations of PM2.5 were 96.5 microg m(-3) and 106.9 microg m(-3) at CGZ and HU site, respectively. More than 80% of the PM2.5 mass concentrations were explained by carbonaceous species, secondary particles, crustal matters and trace elements at the two sites. Carbonaceous species were the most abundant components, constituting about 45% and 48% of the total PM2.5 mass concentrations at CGZ and THU site, respectively. SO4(2-), NO3- and NH4+ were three major ions, accounting for 37%, 23% and 20%, respectively, of the total mass of inorganic water-soluble ions.

Association and dissociation of protein disulfide isomerase.

Purified protein disulfide isomerase, homogeneous by SDS-PAGE, can be separated into two components by PAGE and by gel filtration. These two components, with the same amino-acid composition as well as N- and C-terminal sequences, are the tetramer and dimer of molecular weight 240 kDa and 120 kDa, respectively. The specific activity of the dimer is twice that of the tetramer. At 4 degrees C and pH 7.5 the purified dimer associates and the tetramer dissociates, both slowly and partially, to form a dimer-tetramer mixture. Treatment with dithiothreitol has only a minor effect on the dissociation of the tetramer indicating that the association is not through disulfide formation between the protomers. By prolonged treatment with 1% Triton X-100 or in strong salt solutions the tetramer dissociates to the dimer, but further dissociation to the monomer can only be effected in SDS or guanidine hydrochloride. These results suggest that apart from hydrogen bonds, hydrophobic forces and ionic interactions are mainly involved in the association of the protomers.

FtsZ rings in mukB mutants with or without the Min system.

The site of cell division in Escherichia coli is defined by formation of the Z ring between the two segregated daughter nucleoids. Positioning of the Z ring, composed of the highly conserved and tubulin-like FtsZ protein, appears to be negatively regulated by both the nucleoid and the oscillating MinCD inhibitor proteins. MukB protein is probably involved in nucleoid condensation, and in the absence of MukB, the negative effect of the nucleoid on Z rings appears to be partially suppressed. In this study, we examined the localization of Z rings in cells lacking both the Min system and MukB. In the Deltamin DeltamukB double null mutant, essentially all nucleoid-free zones, either at the cell poles or at non-polar sites between nucleoids, contained Z rings. However, a significant proportion of Z rings also formed on top of nucleoids. Interestingly, Z ring clusters often formed at gaps between nucleoids, and some of the rings within the clusters were clearly positioned on top of nucleoids. These results provide further evidence that the negative topological effect of nucleoids in cells lacking MukB is partially but not totally suppressed, and that the absence of the Min system allows more promiscuous Z ring formation.

Cardiac effects of the extract and active components of Radix stephaniae tetrandrae. I. Electrically-induced intracellular calcium transient and protein release during the calcium paradox.

The present study was designed to compare the cardiac actions of the extract and individual components, tetrandrine (Tet) and fangchinoline (Fan), of Radix stephaniae tetrandrae (RST). We measured the electrically induced [Ca2+]i transient in single rat ventricular myocytes and protein release following perfusion with a Ca2+ free solution (the Ca2+ paradox) from the isolated perfused rat heart, both of which are known to relate to Ca2+ influx. We found that Tet inhibited both electrically induced [Ca2+]i transient and protein release during the Ca2+ paradox, while Fan had no significant effects. The RST extract containing 9% Tet and 6% Fan by weight also affected the [Ca2+]i transient, and was only slightly, though significantly, less effective/potent than Tet alone. On the other hand, RST extract had a significantly greater inhibitory effect on protein release during the Ca2+ paradox than Tet alone. The observations suggest that the RST extract, which contains a mixture of components, may have more potent effects in the heart than its main active component.

Cardiac effects of the extract and active components of radix stephaniae tetrandrae. II. Myocardial infarct, arrhythmias, coronary arterial flow and heart rate in the isolated perfused rat heart.

The primary purpose of the present study was to compare the cardioprotective effects of the extract from radix stephaniae tetrandrae (RST) and its individual compounds, tetrandrine (Tet) and fanchinoline (Fan). Secondly, we also compared the cardiac effects of the individual compounds and the RST extract with those of verapamil, a classical Ca2+ channel blocker. The Langendorff isolated perfused rat heart preparation was used. Regional ischaemia and reperfusion was employed to induce myocardial infarct and arrhythmia. Infarct, arrhythmia, heart rate and coronary artery flow were determined in hearts treated with vehicle, RST extract, Tet, Fan, or verapamil. It was found that RST extract, of which only 9% was Tet, and Tet alone produced equally potent ameliorating effects on arrhythmia and infarct induced by ischaemia and reperfusion without further inhibiting ischaemia-reduced heart rate and coronary artery flow. Fan had no effects on arrhythmia and infarct induced by ischaemia and reperfusion; but it induced S-T segment elevation and further reduced heart rate and coronary artery flow during ischaemia. Verapamil also ameliorated the effects of ischaemia and reperfusion on arrhythmia and infarct. It should be noted that 1 microM verapamil, that produced comparable effects on infarct and arrhythmia to the RST extract and Tet, further inhibited heart rate during ischaemia. The results indicate that the RST extract produces equally potent cardioprotective and anti-arrhythmic effects as Tet alone. Both RST extract and Tet may be better choices for the treatment of arrhythmia and infarct induced by myocardial ischaemia and reperfusion than the classical Ca2+ channel blocker, verapamil as they do not further reduce heart rate during ischaemia.

Antihypertensive and anti-arrhythmic effects of an extract of Radix Stephaniae Tetrandrae in the rat.

In this study, we determined the effects of an extract of Radix Stephaniae Tetrandrae (RST) on arterial blood pressure and heart weight in deoxycorticosterone acetate-salt (DOCA-salt) hypertensive rats. We also determined the effects of the extract on arrhythmia and infarct induced by myocardial ischaemia and reperfusion in anaesthetized rats. We further compared the effects of the extract with those of tetrandrine, which makes up 7% of the extract and is known to act as a calcium-channel antagonist, and verapamil, a prototype calcium-channel antagonist. Treatment with RST extract returned the arterial blood pressure, cardiac compliance and coronary flow towards normal, and reduced right ventricular hypertrophy in the DOCA-salt hypertensive rat. In the anaesthetized rat, the RST extract reduced arrhythmia and infarct size induced by myocardial ischaemia and reperfusion; the effects were similar to those of tetrandrine and verapamil. The findings indicate that the RST extract acts like a calcium-channel antagonist. It may be used in the treatment of cardiovascular diseases, as are the calcium-channel antagonist and tetrandrine. More interestingly, the effects of the RST extract were of the same potency as tetrandrine. Since only 7% of the extract was tetrandrine, the observation indicates that tetrandrine was not the only component that was responsible for the actions of the extract.

Homotransplantation of adrenal gland.

Eight patients with Addison's disease or Cushing's syndrome received adrenal homograft for 9 times after total adrenalectomy. All patients were followed up for 1 to 5 years. Symptoms and signs were improved without the supplement of steroids. The grafts showed satisfactory endocrinal functions in the recipients, but the grafts were dead in two patients 2 and 4 years after operation respectively. One of the two patients received the second homograft with good result. Single perfusion and ice-storage method of the all-adrenal gland and their clinical results were also presented.

[Construction, expression and activity test of a reshaping single-chain antibody against human CD3].

Monoclonal antibody (McAb) against human CD3 can adjust human body's immune statement in various ways, so that its clinical potential is highly regarded. In order to overcome the immunogenecity related to the murine McAb, this research effort was focused on constructing a reshaping single-chain antibody(scFv) against human CD3 employing antibody engineering. First, the CDRs of the murine McAb against human CD3 OKT3 was transplanted into the light-chain framework regions (FRs) of human McAb LS1 and the heavy-chain FRs of human McAb Nd respectively, spatial conformation was predicted by computer analysis. Then some particular residues were replaced in FRs basing on the result of conformational prediction to draw out the amino acid sequences of the reshaped VL and VH. The genes were chemically synthesize and inserted into an expression vector pROH80 to construct the reshaping scFv. Inducing the expression of reshaping scFv, the products are mainly as inclusion bodies. The reshaping scFv was expressed in another vector pALM. The inclusion bodies were denatured and then renatured by gel filtration. The renatured products were purified by immobilized metal affinity chromatograph (IMAC). Finally, the antigen-binding activily of the reshaping scFv against human CD3 was testified by the Compelitire in hibilory fluorescenceactivated cell sorting (FACS). The competitive inhibition rate is 18%.

[Studies of the genetic polymorphism of Schistosoma japonicum (Chinese mainland strains)].

In order to analyze the interspecies diversity and the extent of diversity among Schistosoma japonicum Chinese mainland strains, the genetic variation on gene level among 6 isolates collected from Jiangxi, Hunan, Hubei, Anhui, Sichuan, Yunnan and a cultured isolate from a laboratory were studied using molecular biological technique. There were only two different bases at position 112 and 143 in 536 bp sequence of 28S rDNA-D2 domain between Anhui and Yunnan isolates, the homology was 99.6%. The result could be explained the reason of why the migration rates of single chain 28S rDNA-D2 domain among the above 7 isolates were the same completely in PCR-SSCP. However, comparing with the sequences of the Philippine isolate of S. japonicum, S. mansoni and S. aematobium, there were 6, 94 and 93 different bases and the homologies were 98.9, 82.5 and 82.7 per cent separately. With 8 restriction endonucleases to analyze the ITS of rDNA obtained by PCR from the 7 isolates, the results showed that only 3 minor bands were different, e.g. 5.3% of total 58 fragments. It was suggested that the ITS of rDNA among 7 isolates were highly conserved. Using randomly amplified polymorphic DNA (RAPD) to analyze the genetic diversity of the genomes of the 7 isolates, the average genetic distance (D) calculated from total 284 amplified fragments was 0.22. The maximum D was 0.30 and existed between Anhui and Yunnan isolates. The minimum D was 0.13 and existed between Sichuan and Yunnan isolates. The clustering analysis of genetic distances showed that the 7 isolates could be gathered in one group. From above three results, it could be considered that the genetic diversity on gene level among S. japonicum Chinese mainland strains was very low.

[Homotransplantation of the adrenal gland by microsurgical technique].

Eight patients with Addison's disease or Cushing's syndrome received adrenal homograft following total adrenalectomy for 9 times. Microsurgical technique was used to anastomose the adrenal vessels. All patients were followed up for 1 to 5 years. Symptoms and signs were all improved although without the supplement of steroids. All grafts play satisfactory endocrinal functions in the recipients, but two patients, lost their grafts at 2 and 4 years after operation respectively. One of them underwent the second homograft and the result was good. The single perfusion and ice-storage method of the alloadrenal gland discovered by us and their clinical results were also reported in this paper.

[Gene cloning, construction and expression of single-chain Fv (scFv) against the membrane protein of Schistosoma japonicum].

OBJECTIVE: To construct single chain antibody specific to membrane protein of Schistosoma japonicum by genetic engineering technique. METHODS: The VH (heavy-chain variable region) and VL(light-chain variable region) genes were amplified by PCR from the genomic DNA of NP11-4 cell line, and sequenced by Sanger's method. The ScFv was constructed in pTHA90 vector using VH and VL genes, then expressed by IPTG. RESULTS: The VH and VL genes were obtained through PCR. The DNA sequences showed that VH and VL were new variable region genes of antibody. They were registered by GenBank. A ScFv gene with (Gly4Ser) 3 intralinker in the pTHA90 vector was successfully constructed. The ScFv was expressed as thioredoxin-fused proteins about 36.2 kDa. CONCLUSION: A specific ScFv against the membrane protein of Schistosoma japonicum was constructed and expressed.

[Cloning and sequence analysis of the light chain variable region gene of monoclonal anti-idiotypic antibody NP30 of Schistosoma japonicum].

OBJECTIVE: To amplify and sequence the light chain of anti-idiotypic monoclonal antibody NP30 of Schistosoma japonicum. METHODS: By comparing the conserved regions at each end of the nucleotide sequences of murine germ-line genes encoding FR1 and FR4 regions of immunoglobulin light chain variable regions, we designed a set of primers for amplification of VL gene. The hybridoma cells secreting anti-idiotypic monoclonal antibody NP30 of Schistosoma japonicum were cultured and their genome DNAs were extracted and used as templates for PCR. The PCR product was then cloned into pUC19 vector. The recombinants were sequenced by Sanger's method. The VL gene was compared with GenBank and published mouse VL genes. RESULTS: The full-length of VL gene was 318 bp. The VL gene was a member of mouse Ig kappa light chain subgroup IV and generated from rearrangement of germ line V and J kappa 4 genes. The VL gene sequence has been registered by GenBank(accession No. AF206720). CONCLUSION: The obtained VL gene was a potentially functional gene of anti-idiotypic monoclonal antibody NP30 of Schistosoma japonicum.