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Biomedical subjects

P J Johnson

Publications and source records attributed to P J Johnson.

At least 19 recordsLinked to original sources
PubMed2000

Quantitative comparison of alpha-fetoprotein and albumin mRNA levels in hepatocellular carcinoma/adenoma, non-tumor liver and blood: implications in cancer detection and monitoring.

Early detection of recurrence is valuable for monitoring hepatocellular carcinoma (HCC) progression. By quantitative reverse transcriptase polymerase chain reaction (RT-PCR), we derived calibration curves for alpha-fetoprotein (afp) and albumin (alb) mRNAs using 40 matched tumors and non-tumor liver tissues from HCC/adenoma patients. We prospectively quantified tumor cells and non-tumor liver cells in 62 patients' blood samples before, during and after surgery. Expression of both mRNAs was heterogeneous (1-10(5)-fold) between tumors and HepG2 cell line. The alb-mRNA levels in non-tumor liver cells were 2-10-fold higher than in tumor cells. The afp-mRNA levels in HCC cells were 30-1000-fold higher than in non-tumor cells. The alb-mRNA level in blood may reflect the number of liver cells, whereas the afp-mRNA level may represent mostly the number of HCC cells. We found different ratios of circulating HCC cells to non-tumor liver cells during the clinical course of patients, in association with the subsequent development of recurrence/metastasis. This approach may prove useful for detecting and monitoring HCC progression.

Adenoma, Liver Cell↗
PubMed2000

CTL control of EBV in nasopharyngeal carcinoma (NPC): EBV-specific CTL responses in the blood and tumors of NPC patients and the antigen-processing function of the tumor cells.

Undifferentiated nasopharyngeal carcinoma (NPC) is latently infected with EBV and expresses a restricted number of viral proteins. Studies in healthy virus carriers have demonstrated that at least some of these proteins can act as targets for HLA class I-restricted CTLs. Therefore we have explored the possibility of a CTL-based therapy for NPC by characterizing EBV-specific CTL responses in 10 newly diagnosed NPC cases and 21 healthy virus carriers from Southeast Asia. Using the autologous EBV-transformed lymphoblastoid cell line, virus-specific CTL were reactivated in vitro from PBMC, cloned, and screened for cytotoxicity against target cells expressing individual EBV proteins from recombinant vaccinia vectors. EBV-specific CTLs were identified in 6 of 10 patients and 14 of 21 controls and mainly targeted the EBV nuclear Ag 3 (EBNA3) family of viral latent proteins. However, in 3 of 10 patients and 11 of 21 controls, CTLs specific for the NPC-associated protein LMP2 were also detected, albeit at low frequency. EBV-specific CTLs were detected in tumor biopsy material obtained from 3 of 6 of the patients, indicating that functional CTL are present at the tumor site, but none was specific for tumor-associated viral proteins. To assess the Ag-presenting function in NPC we studied two NPC-derived cell lines (C15 and c666.1) and demonstrated that both were capable of processing and presenting endogenously synthesized protein to HLA class I-restricted CTL clones. Overall, our data provide a sound theoretical basis for therapeutic strategies that aim to boost or elicit LMP2-specific CTL responses in NPC patients.

Adult↗
PubMed2000

Kinetics of plasma Epstein-Barr virus DNA during radiation therapy for nasopharyngeal carcinoma.

We studied the kinetics of circulating EBV DNA in the plasma of nasopharyngeal carcinoma (NPC) patients. Serial weekly sampling of 10 NPC patients revealed a rapid decline in plasma EBV DNA concentration after treatment. In two subjects, an initial rise in the circulating EBV DNA level was observed immediately after treatment initiation. Plasma EBV DNA levels were monitored daily during the first treatment week in a second cohort of five patients, and the results indicated that an initial rise in plasma EBV DNA concentration could be observed in all subjects during the first treatment week. This observation is consistent with the liberation of EBV DNA after therapy-induced cancer cell death. After this initial rise, plasma EBV DNA concentration was found to decay with a median half-life of 3.8 days (interquartile range, 2.4-4.4 days). Kinetic analysis of circulating tumor-derived DNA during treatment may be a powerful tool for evaluating the in vivo response of NPC and other tumors to antineoplastic treatment and may improve our understanding of the biology of plasma nucleic acids.

Carcinoma, Squamous Cell↗
PubMed2000

Gastric impaction and obstruction of the small intestine associated with persimmon phytobezoar in a horse.

Signs of mild colic, intermittent lethargy, and weight loss of 6 weeks' duration in a 2-year-old Quarter Horse gelding were attributed to persimmon (Diospyros virginiana) phytobezoar formation. Diagnosis of the phytobezoar was facilitated by gastric endoscopy. Signs of gastrointestinal tract obstruction were associated with a large phytobezoar in the lumen of the stomach, gastric ulceration, and obstruction of the small intestine (as a consequence of fragmentation of the primary bezoar). Conservative treatment, using mineral oil and dioctyl sodium sulfosuccinate, was unsuccessful. A celiotomy was performed, and gastric impaction and partial obstruction of the small intestine associated with phytobezoar formation and fragmentation were identified. The horse made a complete recovery following removal of all phytobezoars. Persimmon phytobezoar should be considered in the fall and winter as a possible cause of lethargy, colic, and weight loss in horses allowed access to persimmon fruit.

Animals↗
PubMed2000

Lack of expression for the suppressor PML in human small cell lung carcinoma.

The promyelocytic leukemia (PML) gene, which encodes a transformation and growth suppressor, was first identified at the chromosomal translocation break point t(15;17) in acute promyelocytic leukemia (PML). To determine if the PML gene might be involved in other neoplasias such as lung cancer, PML expression was analyzed by immunohistochemical staining and in situ hybridization. Considerable PML protein expression in the PML-oncogenic domain (POD) structure was found in adenocarcinomas (ADC) and squamous cell carcinomas (SCC) of the lung, but was almost completely absent in all the small cell lung carcinomas (SCLC) examined. In situ hybridization showed that both mRNA and DNA of PML were present in SCLC and in normal lung, suggesting that the decreased protein expression was due to either a defect in translation or protein instability, rather than the consequence of decreased transcription or gene deletion. Double staining showed that PML expression was inversely correlated with the proliferation marker Ki-67 and positively correlated with levels of apoptotic cells in these tumors. To determine if the precursor cells of SCLC, the neuroendocrine-producing cells, express PML, double labeling was performed with PML and chromogranin A, a bio-marker for neuroendocrine cells. Neuroendocrine cells from normal tissues were found to be PML positive, indicating that the lack of PML protein in SCLC is associated with the tumorigenic phenotype and is not the result of cell-lineage specificity. Thus, the decreased PML expression may play an important role in SCLC development.

Adenocarcinoma↗
PubMed2000

Competition and protease sensitivity assays provide evidence for the existence of a hydrogenosomal protein import machinery in Trichomonas vaginalis.

Hydrogenosomes are double membrane bounded redox organelles found in a number of amitochondriate protists and fungi. They are involved in carbohydrate metabolism and ATP synthesis and thus resemble mitochondria. Molecular analysis of the hydrogenosomal heat shock proteins Hsp70, Hsp60 and Hsp10 in Trichomonas vaginalis, one of the deepest-branching eukaryotes known to date, has revealed that these group exclusively with mitochondrial heat shock proteins. This finding indicates strongly that a progenitor organelle which gave rise to contemporary mitochondria and hydrogenosomes existed early in eukaryotic life. This hypothesis is further supported by similarities of hydrogenosomal and mitochondrial biogenesis. It was shown that T. vaginalis hydrogenosomal proteins are synthesized on free ribosomes in the cytosol with an N-terminal presequence that carries targeting information and is cleaved upon import into the organelle. Furthermore, as in mitochondrial import, hydrogenosomal protein import requires ATP, an electrochemical transmembrane potential and cytosolic protein factor(s). Here we demonstrate that inhibition of hydrogenosomal protein import occurs (i) in the presence of a synthetic presequence peptide and (ii) after pretreatment of hydrogenosomes with the protease trypsin. Trypsin pretreatment affects two hydrogenosomal membrane proteins of 31 and 70 kDa, respectively. Thus, we present evidence that import is saturable and that proteinaceous hydrogenosomal import receptor(s) exist. These results are a first step towards a characterization of the hydrogenosomal import machinery which should provide further insights into the relationship of hydrogenosomes and mitochondria and the evolution of protein targeting into organelles of endosymbiotic origin.

Animals↗
PubMed2000

Frequent c-myc and Int-2 overrepresentations in nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC) is a commonly occurring tumor in southern China. Although several causative factors have now been recognized, the genetic basis underlying its tumorigenesis is still unclear. To identify potential chromosomal aberrations for further investigations, comparative genomic hybridization (CGH) analysis was applied to the study of genomic imbalances in 10 NPC biopsy specimens. Before CGH analysis, the tumor cell content within the biopsy specimens was enriched by tissue microdissection, and universal genome amplification was performed on the recovered DNA. Recurrent chromosomal gains were detected on 1q (6 of 10 cases), 2q (5 of 10 cases), 3q (7 of 10 cases), 6p (8 of 10 cases), 6q (5 of 10 cases), 7q11.2 (4 of 10 cases), 8q (6 of 10 cases), 11q13, 12, and 15q (8 of 10 cases each), 17q (6 of 10 cases), and 20q (5 of 10 cases). Common losses were identified on 3p (5 of 10 cases), 9p (5 of 10 cases), 11q14-qter (8 of 10 cases), and 14q (5 of 10 cases). Among these aberrations, 7, 8, and 11 gains were further investigated on a series of NPC tissue samples, by interphase fluorescent in situ hybridization (FISH), for the incidence of alpha-satellites: 7, 8, and 11 c-myc and Int-2. Low-level increases of alpha-satellite 7 (9 of 34 cases; 26.5%), alpha-satellite 8 (15 of 34 cases; 44%), and alpha-satellite 11 (8 of 32 cases; 25%) were detected, whereas high-level copy gains of c-myc (21 of 34 cases; 62%) and Int-2 (26 of 34 cases; 76.5%) were more frequently found. Our series is the first to identify genomic overrepresentations of c-myc and Int-2 in NPC. The high incidence of Int-2 amplifications strongly suggests a role of this proto-oncogene in the pathogenesis of NPC.

Adult↗
PubMed2000

TT virus: a study of molecular epidemiology and transmission of genotypes 1, 2 and 3.

BACKGROUND: TT virus (TTV) is a recently discovered virus, which is not related to any other known virus infecting humans. OBJECTIVES: To investigate: (i) the world-wide distribution of the three major TTV genotypes; and (ii) the possible routes of viral transmission. STUDY DESIGN: (i) The phylogenetic distribution of 494 TTV isolates originating from 31 countries was analysed, using partial ORF1 sequences. (ii) Faeces samples (n=22) and saliva samples (n=72) from French individuals were tested for the presence of TTV DNA. (iii) Viral titres in paired serum and saliva samples were compared. RESULTS: (i) Genotypes 1, 2 and 3 were distributed world-wide, with a high proportion of type 1 in Asia (71%) and no type 3 identified in Africa to date. In the USA, 77% of isolates were grouped in four clusters only (genetic distances <10%). This was also the case of 76% of French isolates, 76% of Japanese isolates, and 89% of Hong Kong isolates. (ii) TTV DNA was detected in 18% of faeces samples and 68% of saliva samples tested. (iii) Viral titre in saliva samples was 100-1000 times higher than that of the corresponding serum. CONCLUSIONS: (i) The observed epidemiological distribution of TTV isolates is compatible with an ancient dissemination of viral ancestors belonging to the different genotypes and a slow genetic evolution in sedentary populations. (ii) Besides the possible transmission of TTV by the parental and oral-faecal routes, the high titre of TTV DNA observed in saliva raises the hypothesis of the viral transmission by saliva droplets. This route of transmission could explain the high degree of exposure to viral infection observed in the general population.

Blood Donors↗
PubMed2000

Frequent integration of precore/core mutants of hepatitis B virus in human hepatocellular carcinoma tissues.

The development of hepatitis B virus (HBV)-associated hepatocellular carcinoma (HCC) frequently follows persistent HBV infection and may arise in individuals who are hepatitis B e antigen (HBeAg) negative, indicating the possible presence of precore/core mutants. It is unclear whether precore/core mutants are associated with tumour development or are selected for after chromosomal integration of the wild-type viral DNA. We studied the status and sequence variation of the precore/core region of HBV in 56 patients with HBV-associated HCC and in various corresponding non-tumour tissues by Southern blot analysis, polymerase chain reaction and direct sequencing. Southern blot showed that integrated HBV DNA existed in 43 of 56 HCC tissues. Sequence analysis revealed mutations in 65% of the HCC (26/40) and 45% (14/31) of the corresponding non-tumour tissues. The mutation at nucleotide (nt) 1896, known to prevent HBeAg synthesis, was detected in 40% (16/40) of the tumours and in 35.4% (11/31) of the non-tumour tissues. Other mutations were found at nt 1899 (eight of 40 in HCC; three of 31 in non-tumour tissues), nt 1898 (seven of 40 in HCC; two of 31 in non-tumour tissues), nt 1912 (seven of 40 in HCC; none of 31 in non-tumour tissues) and nt 1886 (three of 40 in HCC; none of 31 in non-tumour tissues). To determine whether this finding merely reflected the prevalence of such mutants in this geographical region, HBV DNA from the sera of patients (also in this region) with acute and chronic hepatitis were sequenced. The nt 1896 mutant was found in 5.6% (one of 18) of patients with acute hepatitis B and in 22.8% (nine of 35) of patients with chronic hepatitis B. However, the nt 1898 mutation was not found in any of these sera. The precore/core mutant was observed with increasing frequency from acute hepatitis to chronic hepatitis, non-tumour and HCC, and this difference in frequency was significant between HCC and acute hepatitis B groups (P < 0.01), suggesting that the precore/core mutant or hepatocytes harbouring this mutant may be under immune selection and that such mutations may facilitate integration and subsequent tumour development.

Adult↗
PubMed2000

Chromosomal translocations are common in natural killer-cell lymphoma/leukemia as shown by spectral karyotyping.

Natural killer (NK)-cell lymphoma/leukemia is a group of rare but highly aggressive neoplasms. The associated genetic aberrations, as defined by conventional cytogenetics, include 6q deletion and chromosome X copy gain, while translocations have been suggested to be uncommon. In this study, three cases of NK cell lymphoma/ leukemia were investigated by spectral karyotyping (SKY). SKY permitted reinterpretation of the chromosomal alterations defined by G-banding and identified several cryptic translocations. In agreement with G-band, 6q deletion was detected in all 3 cases. Structural rearrangement involving chromosome X was observed in 2 cases, and fluorescence in situ hybridization (FISH) analysis indicated that both translocations involved Xp21-pter. Chromosome 8 translocation was also identified in 2 cases and shared a common breakpoint, 8p23. The present study shows the value of SKY in providing additional information on karyotypic abnormalities. The novel findings of recurring Xp21-pter rearrangements and 8p23 translocation should provide basis for further investigations into the tumorigenesis of NK cell lymphoma/leukemia.

Chromosome Banding↗
PubMed2000

Novel intertypic recombinants of epstein-barr virus in the chinese population.

Among 34 Epstein-Barr virus isolates from nonimmunocompromised Chinese donors, we identified three intertypic recombinants with type 1 sequences at the EBNA2 locus and type 2 sequences at some or all of the EBNA3A, -3B, and -3C loci. These appear to have arisen from independent, evolutionarily recent recombination events; such events may be commoner in nonimmunocompromised populations than hitherto imagined.

Base Sequence↗
PubMed2000

Presence of a member of the mitochondrial carrier family in hydrogenosomes: conservation of membrane-targeting pathways between hydrogenosomes and mitochondria.

A number of microaerophilic eukaryotes lack mitochondria but possess another organelle involved in energy metabolism, the hydrogenosome. Limited phylogenetic analyses of nuclear genes support a common origin for these two organelles. We have identified a protein of the mitochondrial carrier family in the hydrogenosome of Trichomonas vaginalis and have shown that this protein, Hmp31, is phylogenetically related to the mitochondrial ADP-ATP carrier (AAC). We demonstrate that the hydrogenosomal AAC can be targeted to the inner membrane of mitochondria isolated from Saccharomyces cerevisiae through the Tim9-Tim10 import pathway used for the assembly of mitochondrial carrier proteins. Conversely, yeast mitochondrial AAC can be targeted into the membranes of hydrogenosomes. The hydrogenosomal AAC contains a cleavable, N-terminal presequence; however, this sequence is not necessary for targeting the protein to the organelle. These data indicate that the membrane-targeting signal(s) for hydrogenosomal AAC is internal, similar to that found for mitochondrial carrier proteins. Our findings indicate that the membrane carriers and membrane protein-targeting machinery of hydrogenosomes and mitochondria have a common evolutionary origin. Together, they provide strong evidence that a single endosymbiont evolved into a progenitor organelle in early eukaryotic cells that ultimately give rise to these two distinct organelles and support the hydrogen hypothesis for the origin of the eukaryotic cell.

Amino Acid Sequence↗
PubMed2000

Hepatitis B virus reactivation in patients undergoing cytotoxic chemotherapy for solid tumours: precore/core mutations may play an important role.

Reactivation of the hepatitis B virus (HBV) is a rare, but well described complication of cytotoxic chemotherapy that may result in hepatic failure. Patients who are chronic carriers of the HBV and who have a G to A mutation at nucleotide 1896 in the precore region may develop more severe liver disease, possibly because of rapid selection and enhanced replication ability of the mutant strain. Such mutant viruses have been implicated occasionally in chemotherapy induced reactivation of hepatitis B virus. In this report, 5 patients with solid tumours were identified to have developed severe hepatitis B virus related liver disease during treatment with cytotoxic agents (with dexamethasone as anti-emetic). All had clinical and serological evidence of reactivation of the HBV. Three patients developed icteric hepatitis; 2 fully recovered, and 1 had died from progressive metastatic disease while recovering from the reactivation. The other two died from progressive liver failure. Direct sequencing of the polymerase chain reaction (PCR) products of the precore (preC) and precore promoter region of the HBV-DNA was carried out on the patients' serum samples taken during the episode of reactivation. In each case, similar mutations (G to A) in nucleotide 1896 of the preC region were found, together with additional mutations in the preC promoter. The present findings suggest that reactivation involving a mutant hepatitis B virus may lead to liver failure, which is possibly more severe than that caused by wild type HBV, and can be triggered by cytotoxic chemotherapy, or the administration of corticosteroids. In Eastern Asia the HBV carriage rate in adults is high. HBV reactivation and severe liver disease during cytotoxic treatment may become a serious and common problem in this region as cytotoxic chemotherapy is more widely used. Patients should be screened routinely for HBsAg in endemic areas of chronic hepatitis B virus infection prior to receiving cytotoxic treatment. The possibility of HBV reactivation should be considered in patients developing liver dysfunction. Patients who are HBeAg negative/Anti-HBe positive, and are suspected to be having an HBV reactivation, should have HBV-DNA levels measured for confirmation as they may carry a mutant HBV.

Adult↗
PubMed2000

Modelling the influence of age, body size and sex on maximum oxygen uptake in older humans.

The purpose of this study was to describe the influence of body size and sex on the decline in maximum oxygen uptake (O2,max) in older men and women. A stratified random sample of 152 men and 146 women, aged 55-86 years, was drawn from the study population. Influence of age on O2,max, independent of differences in body mass (BM) or fat-free mass (FFM), was investigated using the following allometric model: O2,max = BMb (or FFMb) exp(a + (c ' age) + (d ' sex)) [epsilon]. The model was linearised and parameters identified using standard multiple regression. The BM model explained 68.8 % of the variance in O2,max. The parameters (+/- s.e.e., standard error of the estimate) for lnBM (0.563 +/- 0.070), age (-0.0154 +/- 0.0012), sex (0.242 +/- 0.024) and the intercept (-1.09 +/- 0.32) were all significant (P < 0.001). The FFM model explained 69.3 % of the variance in O2,max, and the parameters (+/- s.e.e) lnFFM (0.772 +/- 0.090), age (-0.0159 +/- 0.0012) and the intercept (-1.57 +/- 0.36) were significant (P < 0.001), while sex (0.077 +/- 0.038) was significant at P = 0.0497. Regardless of the model used, the age-associated decline was similar, with a relative decline of 15 % per decade (0.984 exp(age)) in O2,max in older humans being estimated. The study has demonstrated that, for a randomly drawn sample, the age-related loss in O2,max is determined, in part, by the loss of fat-free body mass. When this factor is accounted for, the loss of O2,max across age is similar in older men and women.

Aged↗
PubMed2000

Quantitative analysis of circulating tumor cells in peripheral blood of osteosarcoma patients using osteoblast-specific messenger RNA markers: a pilot study.

Metastasis is a major cause of mortality and morbidity in osteosarcoma (OS) patients. To monitor tumor dissemination, we assessed the circulating tumor burden in OS patients by semiquantitative reverse transcription-PCR using osteocalcin, osteonectin, osteopontin, and type I collagen (COLL) mRNAs as molecular markers. We distinguished levels of the mRNAs in peripheral blood between OS patients and healthy subjects using an OS-derived cell line (Saos-2) as a reference standard. We prospectively analyzed 40 peripheral blood samples from 11 OS patients at diagnosis and 29 healthy subjects. In all 29 (100%) healthy subjects, we detected osteocalcin, osteonectin, and osteopontin mRNAs that were most likely attributed to illegitimate transcription in normal hematopoietic cells. In contrast, we found low COLL mRNA levels in only 35% (10 of 29) of healthy subjects, but significantly higher COLL mRNA levels in 91% (10 of 11) of OS patients (P < 0.0001). The reverse transcription-PCR assay for COLL mRNA was sensitive down to the detection of 10 Saos-2 cells among 10(6) normal peripheral blood nucleated cells. The upper limit of COLL mRNA determined among the healthy subjects was found exceeded by six OS patients. The substantially elevated COLL mRNA levels in peripheral blood seemed to originate from circulating malignant cells in these six OS patients, all of whom subsequently developed clinical metastases within 12 months of diagnosis (P = 0.002). Conversely, no metastases were detected in the remaining OS patients with normal COLL mRNA levels. Quantification of COLL mRNA may prove valuable for diagnosing OS micrometastasis and assessing prognosis.

Adolescent↗