Small single perivascular hepatocellular carcinoma: comparisons of radiofrequency ablation and microwave ablation by using propensity score analysis.

OBJECTIVES: We aimed to compare the therapeutic outcomes of radiofrequency ablation (RFA) and microwave ablation (MWA) as first-line therapies in patients with small single perivascular hepatocellular carcinoma (HCC). METHODS: A total of 144 eligible patients with small (≤ 3 cm) single perivascular (proximity to hepatic and portal veins) HCC who underwent RFA (N = 70) or MWA (N = 74) as first-line treatment were included. The overall survival (OS), disease-free survival (DFS), and local tumor progression (LTP) rates between the two ablation modalities were compared. The inverse probability of treatment weighting (IPTW) method was used to reduce selection bias. Subgroup analysis was performed according to the type of hepatic vessels. RESULTS: After a median follow-up time of 38.2 months, there were no significant differences in OS (5-year OS: RFA 77.7% vs. MWA 74.6%; p = 0.600) and DFS (5-year DFS: RFA 24.7% vs. MWA 40.4%; p = 0.570). However, a significantly higher LTP rate was observed in the RFA group than the MWA group (5-year LTP: RFA 24.3% vs. MWA 8.4%; p = 0.030). IPTW-adjusted analyses revealed similar results. The treatment modality (RFA vs. MWA: HR 7.861, 95% CI 1.642-37.635, p = 0.010) was an independent prognostic factor for LTP. We observed a significant interaction effect of ablation modality and type of peritumoral vessel on LTP (p = 0.034). For patients with periportal HCC, the LTP rate was significantly higher in the RFA group than in the MWA group (p = 0.045). However, this difference was not observed in patients with perivenous HCC (p = 0.116). CONCLUSIONS: In patients with a small single periportal HCC, MWA exhibited better tumor control than RFA. KEY POINTS: • Microwave ablation exhibited better local tumor control than radiofrequency ablation for small single periportal hepatocellular carcinoma. • There was a significant interaction between the treatment effect of ablation modality and type of peritumoral vessel on local tumor progression. • The type of peritumoral vessel is vital in choosing ablation modalities for hepatocellular carcinoma.

[Function of luxS gene in sulfurmetabolism of Streptococcus mutans].

OBJECTIVE: To investigate the function of luxS in sulfurmetabolism of Streptococcus mutans (Sm). METHODS: The growth with absorbency (A) of the standards and mutant strains was measured and analyzed in the sulfur-limited defined medium at different periods. The laser scanning confocal microscopy (LSCM) was used to observe and compare the biofilm thickness of the two kinds of strains at different culture conditions. RESULTS: The significant increases in the thickness of mutant strain biofilm and its growth were observed after the addition of cysteine, but did not reach the standards strain levels (P < 0.05). The growth and the biofilm thickness of the mutant strains were (1.301 ± 0.009) and (45.009 ± 0.429) µm. When methionine and S-adenosylhomocysteine of certain concentrations were respectively added, the biofilm thickness and the growth of mutant strain were raised but did not reach the level of the standards strain at 24 h (P < 0.05), but at 48 h they did. When the methionine was added in the mutant strains for 24 h, the biofilm thickness and the growth of mutant strain were (0.448 ± 0.028) and (37.068 ± 2.392) µm, as for the adding of S-adenosylhomocysteine were (0.460 ± 0.005) and (27.343 ± 1.107) µm. When adding the supernatant fluid of standard strains, the biofilm thickness and the growth levels of mutant strain were much higher than those of the standards strain. The biofilm thickness and growth of both kinds of strains decreased after the addition of S-adenosylmethionine. CONCLUSIONS: luxS gene plays not only a role in quorum sensing but also a role in sulfurmetabolism.

[Application of Cre-loxP(*) system in constructing the markerless double-gene-deletion strain in Streptococcus mutans].

OBJECTIVE: To construct double gene deletions at the htrA and clpP loci on the chromosome of Streptococcus mutans (Sm) and to remove the antibiotic resistance markers with the Cre-loxP(*) site-specific recombination system. METHODS: The htrA gene was cloned into the pGEM-T-Easy TA cloning vector and then inactivated via the insertion of a kanamycin resistance cassette (lox71-Km-lox66), yielding pGEM-T-ΔhtrA/Km for deleting the htrA gene. Using the same method, the pGEM-T-ΔclpP/Sp was constructed for deleting the clpP gene. Following the transformation of pGEM-T-ΔhtrA/Km in Sm, the homologous recombination event was selected. One such mutant was transformed with a cre expression plasmid (pCrePA). The kanamycin resistance gene was then excised. The pCrePA was then easily eliminated at nonpermissive temperatures, resulting in a mutant strain (MSΔhtrA) carrying a deletion at the htrA loci without a selectable marker. This mutant was verified by PCR and DNA sequencing. Then, the clpP and spectinomycin resistance gene were deleted from MSΔhtrA, yielding markerless mutant strain lacking clpP and htrA. RESULTS: The deletion of htrA, clpP and antibiotic resistance markers were confirmed by PCR analysis and DNA sequencing. CONCLUSIONS: A mutant of Sm was constructed successfully which contained a deletion of the htrA and clpP gene without selectable marker. The Cre-loxP(*) system can be applied to Sm, which provides experimental evidence for generating markerless multiple gene deletion mutants.

[Serum soluble Endoglin, plasma endothelin-1 and coagulation function in early onset severe preeclampsia with organ dysfunction].

OBJECTIVE: To investigate the expression levels of serum soluble Endoglin (sEng), plasma endothelin-1 (ET-1) and coagulation function in patients suffering from early onset severe preeclampsia with organ dysfunction, and to analyze the clinical significance. METHODS: Forty-nine early onset severe preeclampsia patients were enrolled in the study group, including 26 cases without organ dysfunction (study group I) and 23 cases with organ dysfunction (study group II). The control group included 30 cases of health pregnant women during the same period of gestation. The serum levels of sEng and plasma ET-1 were analyzed with enzyme-linked immunosorbent assay (ELISA), coagulation function was determined at the same time, and the relationship between the change in levels of sEng, ET-1, coagulation function and organ function, and also outcome of perinatal infants. RESULTS: (1) The levels of sEng, ET-1, fibrinogen (Fib) and mean platelet volume (MPV) of the study group I and II were significantly higher compared with control group (sEng, microg/L: 10.96+/-3.21, 14.17+/-4.02 vs. 7.49+/-2.73; ET-1, microg/L: 41.54+/-10.37, 65.91+/-12.46 vs. 24.56+/-6.26; Fib, g/L: 4.41+/-1.02, 5.35+/-1.17 vs. 3.69+/-0.82; MPV, fl: 11.71+/-1.21, 13.89+/-1.76 vs. 11.03+/-0.82, all P<0.05), and prothrombin time (PT), activated partial thromboplastin time (APTT) and platelet (PLT) were significantly lower compared with control group (PT, s: 10.73+/-1.82, 8.37+/-1.51 vs. 12.95+/-1.91; APTT, s: 26.14+/-4.32, 22.69+/-3.77 vs. 30.25+/-4.71; PLT, x10(9)/L: 164.17+/-50.67, 136.43+/-51.21 vs. 201.63+/-59.83, all P<0.05). There were also statistical significances in all the values between study group I and II (all P<0.05). (2) There was positive correlation between the sEng level and systolic pressure, diastolic pressure, Fib, urine protein of 24 hours, serum creatinine (SCr); there was negative correlation between the sEng level and albumin (Alb) content, PT, estriol/creatinine (E/C) of 12-hour urine, fetal birth weight (all P<0.01). There was positive correlation between the level of ET-1 and the systolic pressure, diastolic pressure, Fib, urine protein of 24 hours, SCr, or alanine aminotransferase (ALT); there was negative correlation between the level of ET-1 and Alb, PT, E/C of 12-hour urine, or fetal birth weight (P<0.05 or P<0.01). (3)In the study group, the occurrence rate of the heart, kidney and lung dysfunction, placental abruption and perinatal death of infants increased (69.23% vs. 11.11%, 38.46% vs. 2.78%, 38.46% vs. 2.78%, 46.15% vs. 2.78%, 53.85% vs. 2.78%, all P<0.01) when the content of sEng>or=16 microg/L compared with sEng<16 microg/L; the occurrence rate of heart, kidney, liver and lung dysfunction, placental abruption and perinatal death of infants increased (64.28% vs. 11.43%, 35.71% vs. 2.86%, 28.57% vs. 5.71%, 28.57% vs. 5.71%, 35.71% vs. 5.71%, 42.86% vs. 5.71%, all P<0.01) when the level of ET-1>or=70 microg/L compared with ET-1<70 microg/L; the occurrence rate of multiple organ dysfunction syndrome was 90% (9/10) when PT<7 s, APTT<20 s and PLT<100x10(9)/L. CONCLUSION: The elevation of levels of serum sEng, plasma ET-1 and coagulation abnormality may contribute to the pathogenesis of the organ dysfunction in early onset severe preeclampsia, and the detection of the above-mentioned indexes has important clinical value.

[Detection of quorum-sensing pathway and construction of luxS gene allelic exchange plasmid of Streptococcus mutans].

OBJECTIVE: To detect the AI-2 quorum-sensing pathway and construct the luxS g-ene allelic exchange plasmid of Streptococcus mutans. METHODS: To detect AI-2 pathway in Streptococcus mutans, the Vibrio harveyi BB170 was used as reporter strain. The PCR fragments of the upstream and downstream regions of luxS and the Erythromycin resistance gene were amplified with the primers respectively, and these fragments were ligated into pUC19 vector with double endonuclease reaction sequentially, the ligated DNAs were transformed into Escherichia coli DH5alpha, then the reconstructed plasmids were isolated and identified by restricted endonuclease digestions. RESULTS: Streptococcus mutans Ingbritt C could induce luminescence of BB170, suggesting the presence of AI-2 quorum sensing pathway in Streptococcus mutans, and such stimulatory activity was maximal at the mid-log growth phase. The recombinant plasmid pUCluxKO was digested by PstI-BamHI, and the digest product were 1000 bp and 5000 bp. When the pUCluxKO was digested by BamHI-KpnI, the digest product were 1500 bp and 4500 bp. While it was digested by KpnI-EcoRI, the digest product were 1000 bp and 5000 bp. All PCR product was in a single belt respectively. CONCLUSIONS: The recombinant plasmid was cloned effectively and can be used in the construction of S.mutans luxS mutant.

[Gene therapy of bladder cancer by using recombinant adeno-associated virus-endostatin: experiments in vitro and in vivo].

OBJECTIVE: To package recombinant adeno-associated virus-endostatin (rAAV-ES) and study its anti-tumor effect in vitro and in vivo. METHODS: rAAV-ES was packaged with co-transfection technique and transfected into the human bladder cancer cells of the line EJ. 24 h later ELISA was used to examine the concentration of ES in the supernatant. The inhibition of human umbilical veins endothelial cells (HUVECs) chemotactic movement were examined by Transwell system. Nude Balb/c mice were divided into 4 groups: (1) 5 mice were inoculated with the EJ cells transfected with rAAV-ES or rAAV-enhanced yellow fluorescence protein (rAAV-EYFP) for 3 days to the subcutaneous tissues of bilateral shoulders so as to observe the growth of tumor. (2) 24 mice were injected with rAAV-ES intramuscularly and then the serum ES was examined every 10 days since the 10 th day after the injection. (3) 36 mice were randomly subdivided into 3 equal subgroups to be injected with rAAV-ES, rAAV-EYFP, or RPMI medium, inoculated with EJ cells 2 weeks later, and then killed 50 days later to observe the size of tumor. (4) 4 healthy mice and 4 mice injected with rAAV-ES for 8 weeks were killed with their hearts and brains taken out to observe the side effects. RESULTS: rAAV-ES was packaged successfully. The ES concentration in the supernatant of culture fluid of the EJ cell transfected with rAAV-ES was 54.09 ng/ml. The inhibition rate of the HUVECs chemotactic movement was 37.45%. The xenograft formation rate was 2/5 for the EJ cells transfected with rAAV-ES. The serum ES levels of the mice injected with rAAV-ES remained high. The tumor size in the mice injected with rAAV-ES was significantly smaller than those of the other groups (both P < 0.01). No pathological changes was found in the hearts and brains in the mice injected with rAAV-ES. CONCLUSION: rAAV-ES inhibits tumor angiogenesis, and tumor formation and progression. Successful packaging of rAAV-ES has laid a foundation for gene therapy of bladder cancer.