OBJECTIVETo assess the level of alpha-fetoprotein (AFP) messenger RNA (mRNA) in peripheral blood of nude mice, and to study its relationship with tumor recurrence and metastasis after curative resection of hepatocellular carcinoma (HCC).

METHODSThe metastatic model of human HCC in nude mice LCI-D20 was used in this study. Curative resection was performed at 10th day after tumor implantation in 28 nude mice. Interferon alpha-1b (IFN alpha-1b) was administered subcutaneously from the next day after resection, at doses of 3 10(7)U/kg/d (8 nude mice), 1.5 10(7) U/kg/d (8 nude mice) respectively in the treatment groups, and normal saline alone in the controlled group (12 nude mice). Thirty-five days after treatment, one milliliter of peripheral blood was taken and AFP mRNA was quantitatively analyzed using TaqMan real-time quantitative RT-PCR. The mice were sacrificed. The size of recurrent tumor was measured and the presence of lung metastases was observed.

RESULTSThe liver recurrent rate, lung metastatic rate and positivity of AFP mRNA in the controlled group were all 100% (12/12), whereas it was 62.5% (5/8), 0% (0/8), 87.5% (7/8) respectively in the IFN alpha-1b 1.5 10(7)U/kg/d treated group. The recurrent tumor in liver of the IFN alpha-1b 1.5 10(7)U/kg/d treated group was much smaller than that of the controlled group (25 mm(3) 2 mm(3) vs 1143 mm(3) 3 mm(3), t =9.27, P<0.01), and the level of AFP mRNA was also lower than that of the controlled group [(85 6)copies/mug vs (955 2) copies/mug, t =4.33, P<0.01]. In the IFN alpha-1b 3 10(7)U/kg/d treated group, there was only one recurrent tumor (0.5 mm(3)), no lung metastasis, and the positivity of AFP mRNA was 0% (?(2)=11.67, P<0.01).

CONCLUSIONSThese results suggest that the level of AFP mRNA in peripheral blood may indicate recurrence and/or metastasis after curative resection of HCC. TaqMan real time quantitative RT-PCR is a very sensitive and convenient method for detecting circulating cancer cells.

Animals ; Biomarkers, Tumor ; analysis ; genetics ; Carcinoma, Hepatocellular ; secondary ; surgery ; Disease Models, Animal ; Liver Neoplasms ; pathology ; surgery ; Mice ; Mice, Nude ; Neoplasm Metastasis ; RNA, Messenger ; analysis ; Recurrence ; alpha-Fetoproteins ; analysis ; genetics

OBJECTIVETo clone the unknown gene of hepatocyte protein interacting with hepatitis C virus core protein.

METHODSUsing the yeast dual hybrid system 3, bait plasmids of hepatitis C virus core were constructed. After identifying hepatitis C virus core protein that could stably expressed in AH109 yeast strains, we performed yeast two hybrid by mating AH109 with Y187 that transformed with liver cDNA library plasmids pACT2 and then plated on quadrople dropout (QDO) medium and assayed for alpha-gal activity. The genes of yeast colonies that could grow on QDO and had alpha-gal activity were sequenced.

RESULTSAmong the 30 positive colonies, we blasted the gene of the sixth colony; we coined human hepatitis C virus binding protein 6(Hu Hcbp6) with Genbank, realized that the Hu Hcbp6 shares as much as 98% homology with two cDNA without knowing functions. We have proved that Hu Hcbp6 could interact with hepatitis C virus core protein.

CONCLUSIONSHepatitis C virus core binding protein (Hu Hcbp 6 Genbank number: AY032594) was successfully cloned and identified. The study partly paved the way for investigating physiological function of the Hu Hcbp6.

Cloning, Molecular ; DNA, Complementary ; genetics ; DNA-Binding Proteins ; genetics ; Hepacivirus ; Humans ; Molecular Sequence Data ; Plasmids ; Sequence Analysis, DNA ; Transfection ; Two-Hybrid System Techniques ; Viral Core Proteins ; genetics ; metabolism ; Yeasts ; genetics

This review deals with the characteristics of various inflammatory mediators identified in the middle ear during otitis media and in cholesteatoma. The role of each inflammatory mediator in the pathogenesis of otitis media and cholesteatoma has been discussed. Further, the relation of each inflammatory mediator to the pathophysiology of the middle and inner ear along with its mechanisms of pathological change has been described. The mechanisms of hearing loss including sensorineural hearing loss (SNHL) as a sequela of otitis media are also discussed. The passage of inflammatory mediators through the round window membrane into the scala tympani is indicated. In an experimental animal model, an application of cytokines and lipopolysaccharide (LPS), a bacterial toxin, on the round window membrane induced sensorineural hearing loss as identified through auditory brainstem response threshold shifts. An increase in permeability of the blood-labyrinth barrier (BLB) was observed following application of these inflammatory mediators and LPS. The leakage of the blood components into the lateral wall of the cochlea through an increase in BLB permeability appears to be related to the sensorineural hearing loss by hindering K+ recycling through the lateral wall disrupting the ion homeostasis of the endolymph. Further studies on the roles of various inflammatory mediators and bacterial toxins in inducing the sensorineumral hearing loss in otitis media should be pursued.
Bacterial Toxins ; Chemokines ; Cholesteatoma ; Cochlea ; Cytokines ; Ear, Inner ; Ear, Middle ; Endolymph ; Evoked Potentials, Auditory, Brain Stem ; Hearing Loss ; Hearing Loss, Sensorineural ; Homeostasis ; Membranes ; Models, Animal ; Otitis ; Otitis Media ; Permeability ; Recycling ; Scala Tympani