Monocarboxylate transporter 1-mediated lactate accumulation promotes nucleus pulposus degeneration under hypoxia in a 3D multilayered nucleus pulposus degeneration model.

During intervertebral disc degeneration (IVDD), due to endplate calcification, diminished oxygen and nutrient concentrations and accumulated lactate are present in the microenvironment of the nucleus pulposus (NP). The disadvantages of 3D layered culture include uneven oxygen and nutrient gradients. In the present study, to mimic the in vivo microenvironment of the NP, a 5-layered 3D culture was constructed using clinical haemostatic gelatine sponges and developed as a NP degeneration (NPD) model. Subsequently, cell distribution as well as expression of NP chondrogenic markers (type II collagen and aggrecan), glycosaminoglycan (GAG) and degeneration markers [e.g. matrix metalloproteinase (MMP) 3] were measured from the top to the bottom layer. However, in a single NP-cell-loaded disc model, the chondrogenic potency in the middle or bottom layer was higher than that in the top layer. To further study the mechanism underlying the degeneration of NP cells in this NPD model, the contribution of secreted metabolites was examined. Lactate identified in the supernatant modulated GAG accumulation and MMP3 expression. Inhibition of lactate influx by the monocarboxylate transporter (MCT)-1 inhibitor, AZD3965, reversed the effect of lactate on GAG accumulation and MMP3 expression and further improved NP cell degeneration in the NPD model. Thanks to the homogenous expression of lactate in the model, it was possible to further identified that the combination of lactate and hypoxia enhanced MMP3 expression. Taken together, multilayered cell-loaded sponges, with oxygen and nutrient gradients as well as lactate accumulation, can represent a 3D multilayered NPD model for exploring potential agents for IVDD.

Overview of Japanese encephalitis disease and its prevention. Focus on IC51 vaccine (IXIARO®).

Japanese encephalitis (JE) is a vector-borne disease caused by the Japanese encephalitis virus (JEV). JEV is transmitted by mosquitoes to a wide range of vertebrate hosts, including birds and mammals. Domestic animals, especially pigs, are generally implicated as reservoirs of the virus, while humans are not part of the natural transmission cycle and cannot pass the virus to other hosts. Although JEV infection is very common in endemic areas (many countries in Asia), less than 1% of people affected develop clinical disease, and severe disease affects about 1 case per 250 JEV infections. Although rare, severe disease can be devastating; among the 30,000-50,000 global cases per year, approximately 20-30% of patients die and 30-50% of survivors develop significant neurological sequelae. JE is a significant public health problem for residents in endemic areas and may constitute a substantial risk for travelers to these areas. The epidemiology of JE and its risk to travelers have changed, and continue to evolve. The rapid economic growth of Asian countries has led to a surge in both inbound and outbound travel, making Asia the second most-visited region in the world after Europe, with 279 million international travelers in 2015. The top destination is China, followed by Thailand, Hong Kong, Malaysia and Japan, and the number of travelers is forecast to reach 535 million by 2030 (+ 4.9% per year). Because of the lack of treatment and the infeasibility of eliminating the vector, vaccination is recognized as the most efficacious means of preventing JE. The IC51 vaccine (IXIARO®) is a purified, inactivated, whole virus vaccine against JE. It is safe, well tolerated, efficacious and can be administered to children, adults and the elderly. The vaccination schedule involves administering 2 doses four weeks apart. For adults, a rapid schedule (0-7 days) is available, which could greatly enhance the feasibility of its use. Healthcare workers should inform both short- and long-term travelers of the risk of JE in each period of the year and recommend vaccination. Indeed, it has been shown that short-term travelers are also at risk, not only in rural environments, but also in cities and coastal towns, especially in tourist localities where excursions to country areas are organized.

Influenza vaccination of elderly: relaunch time.

Influenza can be a serious disease and constitutes a threat to the population. Every year, seasonal influenza epidemics affect about 5-15% of the world's population. Some frail categories (such as the elderly) can develop complications, request hospitalization, and may die. In order to reduce the medical, social and economic burden of influenza, vaccination is recommended by many health authorities worldwide. Italy has a national programme of influenza vaccination which targets specific categories, such as subjects with chronic conditions, pregnant women, healthcare workers and those over 65 years old. Despite this opportunity for prevention, however, vaccination coverage in Italy does not reach the minimum recommended threshold of 75%. This paper reports some interventions that can improve coverage rates of the elderly, such as "tailor-made" information campaigns, healthcare workers training and the adoption of innovative communication strategies in order to implement vaccination strategies that take into account the needs of the elderly population, the involvement of elderly people's associations in awareness-raising activities and strengthening the role of general practitioners in promoting influenza vaccination.

Influenza vaccination: from epidemiological aspects and advances in research to dissent and vaccination policies.

Influenza is a serious public health problem, since seasonal epidemics affect approximately 5-10% of the population and thus give rise to a heavy social and healthcare burden. The heavy burden of disease is due to several factors, one of which is the biological features of the pathogen. Indeed influenza viruses display high mutation rates and undergo frequent genetic reassortment. Minor variations cause seasonal epidemics and major variations, which result from the hybridization of viruses typical of different animal species, can lead to pandemics. Vaccination remains the most efficacious means of mitigating the harmful healthcare and social effects of influenza. Influenza vaccines have evolved over time in order to offer broader protection against circulating strains. Trivalent vaccines containing two A viruses and one B virus are currently available. However, given the co-circulation of both B virus lineages (B/Yamagata and B/Victoria), quadrivalent vaccines have recently been developed. The new quadrivalent vaccines constitute a great advance, in that they can offer broader strain coverage. Despite the availability of effective and safe influenza vaccines, the Italian public's trust in vaccination has declined and, in the last few years, influenza vaccination coverage rates have decreased both among the elderly and among at-risk adults. It is therefore necessary that users, in their own interests, regain trust in this important means of disease prevention. In order to mitigate the damage wreaked by influenza, it seems important to: (i) improve clinical-epidemiological and virological surveillance of the disease; (ii) promote the development of new efficacious vaccines, as has recently been done through the introduction of the quadrivalent vaccine; (iii) extend free vaccination to the entire population, as in the US and Canada; (iv) ensure that general healthcare professionals are properly informed and always updated with regard to vaccination; (v) promote public campaigns to raise the population's awareness of the importance of vaccination; (vi) inform politicians and other decision-makers of scientific results in the field of vaccination; (vii) fight the antivaccination lobbies with every available weapon.

Meningococcal B vaccination strategies and their practical application in Italy.

Immunisation against meningococcal meningitis has a long history, which has passed through several phases: the studies by Flexner, extraction of the polysaccharide capsule, the development of monovalent and multivalent conjugate vaccines, the outer membrane vesicle vaccines up to the development of effective and safe vaccines for meningococcal B invasive disease through the application of the techniques of molecular biology and reverse vaccinology. The new available vaccines are Bexsero® and Trumenba®. Bexsero ® has been approved and is available in Europe, the USA, Canada, Australia and Chile, and is currently under review in Brazil for the prevention of MenB invasive disease in subjects ≥ 2 months. Trumemba® is currently approved only in the USA, for use in adolescents and young adults. At present, the greatest obstacle to the extensive use of these vaccines in industrialised countries is the high cost and the need administer multiple doses in infants. However, in some European countries and in some Italian Regions, strategies (free and active call) to fight the disease through vaccination (Bexsero®) are already in place.

The "urban myth" of the association between neurological disorders and vaccinations.

In modern society, a potentially serious adverse event attributed to a vaccination is likely to be snapped up by the media, particularly newspapers and television, as it appeals to the emotions of the public. The widespread news of the alleged adverse events of vaccination has helped to create the "urban myth" that vaccines cause serious neurological disorders and has boosted antivaccination associations. This speculation is linked to the fact that the true causes of many neurological diseases are largely unknown. The relationship between vaccinations and the onset of serious neuropsychiatric diseases is certainly one of coincidence rather than causality. This claim results from controlled studies that have excluded the association between vaccines and severe neurological diseases, therefore it can be said, with little risk of error, that the association between modern vaccinations and serious neurological disorders is a true "urban myth".

Compounds with anti-influenza activity: present and future of strategies for the optimal treatment and management of influenza. Part I: Influenza life-cycle and currently available drugs.

Influenza is a contagious respiratory acute viral disease characterized by a short incubation period, high fever and respiratory and systemic symptoms. The burden of influenza is very heavy. Indeed, the World Health Organization (WHO) estimates that annual epidemics affect 5-15% of the world's population, causing up to 4-5 million severe cases and from 250,000 to 500,000 deaths. In order to design anti-influenza molecules and compounds, it is important to understand the complex replication cycle of the influenza virus. Replication is achieved through various stages. First, the virus must engage the sialic acid receptors present on the free surface of the cells of the respiratory tract. The virus can then enter the cells by different routes (clathrin-mediated endocytosis or CME, caveolae-dependent endocytosis or CDE, clathrin-caveolae-independent endocytosis, or macropinocytosis). CME is the most usual pathway; the virus is internalized into an endosomal compartment, from which it must emerge in order to release its nucleic acid into the cytosol. The ribonucleoprotein must then reach the nucleus in order to begin the process of translation of its genes and to transcribe and replicate its nucleic acid. Subsequently, the RNA segments, surrounded by the nucleoproteins, must migrate to the cell membrane in order to enable viral assembly. Finally, the virus must be freed to invade other cells of the respiratory tract. All this is achieved through a synchronized action of molecules that perform multiple enzymatic and catalytic reactions, currently known only in part, and for which many inhibitory or competitive molecules have been studied. Some of these studies have led to the development of drugs that have been approved, such as Amantadine, Rimantadine, Oseltamivir, Zanamivir, Peramivir, Laninamivir, Ribavirin and Arbidol. This review focuses on the influenza life-cycle and on the currently available drugs, while potential antiviral compounds for the prevention and treatment of influenza are considered in the subsequent review.

Compounds with anti-influenza activity: present and future of strategies for the optimal treatment and management of influenza. Part II: Future compounds against influenza virus.

In the first part of this overview, we described the life cycle of the influenza virus and the pharmacological action of the currently available drugs. This second part provides an overview of the molecular mechanisms and targets of still-experimental drugs for the treatment and management of influenza. Briefly, we can distinguish between compounds with anti-influenza activity that target influenza virus proteins or genes, and molecules that target host components that are essential for viral replication and propagation. These latter compounds have been developed quite recently. Among the first group, we will focus especially on hemagglutinin, M2 channel and neuraminidase inhibitors. The second group of compounds may pave the way for personalized treatment and influenza management. Combination therapies are also discussed. In recent decades, few antiviral molecules against influenza virus infections have been available; this has conditioned their use during human and animal outbreaks. Indeed, during seasonal and pandemic outbreaks, antiviral drugs have usually been administered in mono-therapy and, sometimes, in an uncontrolled manner to farm animals. This has led to the emergence of viral strains displaying resistance, especially to compounds of the amantadane family. For this reason, it is particularly important to develop new antiviral drugs against influenza viruses. Indeed, although vaccination is the most powerful means of mitigating the effects of influenza epidemics, antiviral drugs can be very useful, particularly in delaying the spread of new pandemic viruses, thereby enabling manufacturers to prepare large quantities of pandemic vaccine. In addition, antiviral drugs are particularly valuable in complicated cases of influenza, especially in hospitalized patients. To write this overview, we mined various databases, including Embase, PubChem, DrugBank and Chemical Abstracts Service, and patent repositories.

New versus old meningococcal group B vaccines: how the new ones may benefit infants & toddlers.

Invasive disease caused by Neisseria meningitidis is associated with high mortality and high disability rates and mainly affects children under one year of age. Vaccination is the best way to prevent meningococcal disease, especially in infants and toddlers. The introduction of massive meningococcal serogroup C vaccination has drastically reduced the incidence of disease caused by this serogroup, and serogroup B has now become the main causative agent in several industrialized countries. The first serogroup B vaccines, which were used for more than two decades, were based on outer membrane vesicles and proved to be protective only against specific epidemic strains in Cuba, Norway, Brazil and New Zealand. Moreover, these often elicited a scant immune response in young children. Innovative genomics-based reverse vaccinology subsequently enabled researchers to identify genes encoding for surface proteins that are able to elicit a strong immune response against several B strains. This important discovery led to the development and recent approval in Europe of the four-component meningococcal serogroup B (4CMenB) vaccine. Large clinical trials have shown high immunogenicity and tolerability and acceptable safety levels of 4CMenB in infants and toddlers. This vaccine is expected to cover a large number of circulating invasive strains and may also be efficacious against other serogroups. Young children are particularly vulnerable to the devastating consequences of meningococcal disease. Given the high performance of 4CMenB and its non-interference with routine vaccinations, this age-group will be the first to benefit from the introduction of this vaccine.

Tobacco smoking among students in an urban area in Northern Italy.

INTRODUCTION: Tobacco smoking, which usually begins in teenage, is one of the most important lifestyle risk factors for chronic diseases and a major public health problem worldwide. The aims of the study were to determine the prevalence of tobacco smoking and the mean age of initiation among adolescents in Genoa (Italy) and to identify some socio-demographic predictors that could be associated with the onset of smoking. MATERIALS AND METHODS: 2,301 randomly selected students (14-19 years old) in Genoa completed an ad hoc questionnaire. The Kaplan-Meier method was used to evaluate the instantaneous risk of experimenting with smoking. A multivariate logistic regression model was used to determine whether current or previous smoking status was associated with socio-demographic characteristics. RESULTS: 59.5% of respondents had tried smoking, while 35.6% defined themselves as current smokers. No difference in current smoking prevalence emerged between males and females (35.2% and 35.9%, respectively, p = 0.83). The mean age on initiation was 13.5 years for males and 13.9 years for females. The instantaneous probability of trying smoking changed with age, reaching a maximum at 14 years. Subjects who tried smoking before this age were more inclined to continue smoking. The probability of being a current smoker was significantly higher among students from unmarried-parent families and those attending vocational and technical secondary schools. CONCLUSIONS: There is a great need for the activation of new health promotion interventions and enforcement of those already existing, in order to raise awareness of the damage caused by smoking among adolescents, especially those belonging to high-risk groups.

Neisseria meningitidis, pathogenetic mechanisms to overcome the human immune defences.

Neisseria meningitidis is hosted only by humans and colonizes the nasopharynx; it survives in the human body by reaching an equilibrium with its exclusive host. Indeed, while cases of invasive disease are rare, the number of asymptomatic Neisseria meningitides carriers is far higher. The aim of this paper is to summarize the current knowledge of survival strategies of Neisseria meningitides against the human immune defences. Neisseria meningitidis possesses a variety of adaptive characteristics which enable it to avoid being killed by the immune system, such as the capsule, the lipopolysaccharide, groups of proteins that block the action of the antimicrobial proteins (AMP), proteins that inhibit the complement system, and components that prevent both the maturation and the perfect functioning of phagocytes. The main means of adhesion of Neisseria meningitides to the host cells are Pili, constituted by several proteins of whom the most important is Pilin E. Opacity-associated proteins (Opa) and (Opc) are two proteins that make an important contribution to the process of adhesion to the cell. Porins A and B contribute to neisserial adhesion and penetration into the cells, and also inhibit the complement system. Factor H binding protein (fhbp) binds factor H, allowing the bacteria to survive in the blood. Neisserial adhesin A (NadA) is a minor adhesin that is expressed by 50% of the pathogenic strains. NadA is known to be involved in cell adhesion and invasion and in the induction of proinflammatory cytokines. Neisserial heparin binding antigen (NHBA) binds heparin, thus increasing the resistance of the bacterium in the serum.

Live attenuated influenza vaccine--a review.

Owing to the variability of influenza viruses, vaccine composition needs to be up-dated annually. As many variables can influence their efficacy, vaccines are still considered "sub-optimal". Many studies have been carried out in recent years to improve vaccines. In particular, researchers and vaccine-producing corporations have focused on developing a live vaccine. Among the candidate vaccines, the strain developed by Maassab has recently been licensed in the USA and Europe, after extensive investigation. This vaccine is safe and well tolerated, and has shown very good genetic stability. Although vaccine recipients are able to spread the virus, transmission to close contacts is practically non-existent. Studies on cold-adapted attenuated influenza vaccines have demonstrated that such vaccines are effective, and sometimes more effective than inactivated influenza vaccines. Cold-adapted attenuated influenza vaccines therefore appear to be an important weapon against influenza. However, a more widespread use of these vaccines is to be recommended, especially in children, as the more acceptable way of administration can favour parental compliance.

Utility of thymosin alpha-1 (Zadaxin) as a co-adjuvant in influenza vaccines: a review.

Influenza constitutes a serious problem for healthcare and social services worldwide, owing to its pattern and the severity of its complications in some categories of subjects at risk, such as the elderly and immunocompromised individuals. The only really effective means of combating influenza is vaccination. The elderly and immunocompromised subjects are refractory or low responders to vaccination. The need for ever more immunogenic and efficacious influenza vaccines, especially for subjects at risk, has prompted the development of adjuvated vaccines. With a view to enhancing the immune response in the elderly and in subjects at risk, the possibility of co-administering immunostimulants as Thymosin alpha-1 (Talpha1) with influenza vaccines has been investigated. Talpha1 is a biologically active peptide made up of 28 amino acids that can enhance T-cells, dendritic cell and antibody responses, modulate cytokines and chemokines production. Several studies were conducted and showed that Talpha1 ameliorate the performanc of influenza vaccination in elderly and subjects at risk. Although further studies on co-adjuvants are necessary, the future prospects of producing ever more efficacious influenza vaccines appear very promising.

[Changes of WT1 mRNA expression level in patients with myelodysplastic syndromes after hypomethylating agents and its prognostic significance].

Objective: To monitor the WT1 mRNA level and its dynamic changes in patients with myelodysplastic syndromes (MDS) after hypomethylating agents (HMA) , as well as to assess the significance of WT1 mRNA levels and its dynamic changes in evaluating the efficacy of HMA and distinguishing the disease status of heterogeneous patients with stable disease (SD) . Methods: Bone marrow or peripheral blood samples of 56 patients with MDS who underwent hypomethylating agents (≥4 cycles) from November 2009 to March 2018 were tested by real-time quantitative polymerase chain reaction (PCR) to detect the expression of WT1 mRNA, and to observe the correlation between the dynamic changes of WT1 mRNA expression and clinical efficacy and prognosis of patients. Results: WT1 mRNA expression levels of MDS patients decreased significantly after 3 cycles of hypomethylating agent treatment. Besides, the WT1 mRNA expression levels of patients increased significantly after diseases progression. According to the dynamic changes of WT1 mRNA expression levels during SD, 45 cases could be further divided into increased group and non-increased group. In those SD patients with increased WT1 mRNA expression level, the ratio of suffering disease progression or transformation to AML was 95.65% (22/23) , whereas the ratio turned to be 9.09% (2/22) for the non-increased group (χ(2)=33.852, P<0.001) . Compared with those SD patients reporting no increase in WT1 mRNA expression level, the overall survival[17 (95%CI 11-23) months vs not reached, P<0.001] and progression-free survival [13 (95%CI 8-18) months vs not reached, P<0.001] of those SD patients reporting increase in WT1 mRNA expression level were significantly shorter. Conclusion: WT1 mRNA expression level is a useful indicator to assess the efficacy of hypomethylating agents in MDS patients. Especially in patients with SD, detection of the changes in WT1 mRNA expression level is able to predict disease progression and help to make clinical decision.

[Artesunate attenuate chronic graft-versus-host disease by regulating Th17/Treg balance].

Objective: To investigate the effects of artesunate treatment on chronic graft-versus-host disease (cGVHD). Methods: Recipient BALB/c mice received 8 × 10(6) bone marrow cells with 8×10(6) spleen cells from B10D2 mice. Artesunate solubilized in acetone was injected intraperitoneally every day at the dose of 1 mg/kg at Day 28 after BMT. The clinical scores, survival and histopathological damage were analyzed. The frequency of Th17 and Tregs in PB and spleens from the mice were evaluated by flow cytometry. In addition, CD4(+) T cells from the spleens of mice were cultured in vitro, then stimulated with artesunate, the frequency of Th17 and Tregs in these splenocytes were evaluated by flow cytometry. Results: Artesunate administration diminished clinical and histopathological damage, and improved the survival of cGVHD mice[(46.57±7.83)% vs (55.71±6.99)%, χ(2)=5.457, P=0.020]; Artesunate contributed to Tregs development [(4.45±0.04)% vs (8.40±0.23)%, t=15.679, P<0.001; (6.62±0.24)% vs (10.48±0.48)%, t=6.587, P=0.003] while decreased Th17 cells [(1.51±0.18)% vs (0.58±0.19)%, t=7.233, P<0.001; (1.48±0.38)% vs (0.71±0.18)%, t=3.653, P=0.011] expressions in both PB and spleens, and decreased the Th17/Treg ratio (0.34±0.05 vs 0.09±0.03, t=7.621, P=0.002; 0.19±0.03 vs 0.06±0.02, t=6.993, P=0.002). Moreover, artesunate suppressed the Th17 cells expressions [(0.82±0.37) % vs (3.39±1.22) %, t=4.044, P=0.007] and contributed to Tregs development [(34.63±1.29) % vs (14.28±1.69) %, t=19.119, P<0.001], and also decreased the Th17/Treg ratio (0.24±0.09 vs 0.02±0.01, t=4.780, P=0.003) in vitro. Conclusions: Artesunate suppressed the Th17 cells expressions and contributed to Tregs development, which provided new sights into the development of a novel drug for cGVHD, e.g., artemisinin.

Burden of typhoid fever and cholera: similarities and differences. Prevention strategies for European travelers to endemic/epidemic areas.

The burden of diarrheal diseases is very high, accounting for 1.7 to 5 billion cases per year worldwide. Typhoid fever (TF) and cholera are potentially life-threatening infectious diseases, and are mainly transmitted through the consumption of food, drink or water that have been contaminated by the feces or urine of subjects excreting the pathogen. TF is mainly caused by Salmonella typhi, whereas cholera is caused by intestinal infection by the toxin-producing bacterium Vibrio cholerae. These diseases typically affect low- and middle-income countries where housing is overcrowded and water and sanitation are poor, or where conflicts or natural disasters have led to the collapse of the water, sanitation and healthcare systems. Mortality is higher in children under 5 years of age. Regarding their geographical distribution, TF has a high incidence in sub-Saharan Africa, India and south-east Asia, while cholera has a high incidence in a few African countries, particularly in the Horn of Africa and the Arabian Peninsula. In the fight against these diseases, preventive measures are fundamental. With modern air travel, transmissible diseases can spread across continents and oceans in a few days, constituting a threat to global public health. Nowadays, people travel for many reasons, such as tourism and business. Several surveys have shown that a high proportion of travelers lack adequate information on safety issues, such as timely vaccination and prophylactic medications. The main objective of this overview is to provide information to help European travelers to stay healthy while abroad, and thus also to reduce the potential importation of these diseases and their consequent implications for public health and society. The preventive measures to be implemented in the case of travel to countries where these diseases are still endemic are well known: the adoption of safe practices and vaccinations. It is important to stress that an effective preventive strategy should be based both on vaccinations and on hygiene travel guidelines. Furthermore, the emergence of multidrug-resistant strains is becoming a serious problem in the clinical treatment of these diseases. For this reason, vaccination is the main solution.

Overexpression of tumour-associated trypsin inhibitor (TATI) enhances tumour growth and is associated with portal vein invasion, early recurrence and a stage-independent prognostic factor of hepatocellular carcinoma.

Tumour-associated trypsin inhibitor (TATI) overexpresses in various tumours, but its clinicopathological significance in hepatocellular carcinoma (HCC) is unclear. Differential display analysis revealed expression of TATI in HCC. By RT-PCR in the linear range, TATI was found to be overexpressed in 176 of 258 unifocal primary HCCs (68%). TATI overexpression correlated with high-stage HCC (stage IIIB to IV) with portal vein (PV) invasion (p=0.00014), early tumour recurrence (ETR; p=0.00002), and a lower 5-year survival (p=0.000001), in both low- and high-stage HCC (p=0.033 and p=0.00036, respectively). Ectopic expression of TATI led to enhanced anchorage-independent tumour cell growth in vitro. To determine its potential as a part of a group of combined diagnostic markers, we analysed 235 HCCs for three genes encoding secretory proteins known to be overexpressed in HCC; these were TATI, AFP and osteopontin; 202 of the tumours (86%) overexpressed one or more of these genes. Further, HCC with a greater number of gene overexpressions produced bigger tumours (p=0.0024), had a higher rate of PV invasion (p= 1x10(-8)), had a higher ETR (p=1x10(-8)), and showed a lower 5-year survival (p=0.000001). We conclude that TATI overexpression contributes to cell growth advantage, enhances the metastatic potential of tumours and leads to advanced HCC with PV invasion. Thus, it is a stage-independent prognostic factor for HCC and a useful predictor for ETR. Moreover, it should be possible to use TATI, AFP and osteopontin as combined markers for molecular staging, the detection of HCC and for the prediction of ETR.

Cloning and expression of a developmentally regulated transcript MXR7 in hepatocellular carcinoma: biological significance and temporospatial distribution.

Using the differential display method to analyze mRNA expression in hepatocellular carcinoma (HCC) and nontumor livers, we cloned a full-length cDNA of 2263 bp, which was designated GTR2-2 and was identical with MXR7. The MXR7 mRNA was detected in 143 of 191 (74.8%) primary and recurrent HCCs taken from 154 patients but only in 5 (3.2%) nontumor livers. MXR7 mRNA was detected in one of two hepatoblastomas but not in hepatocellular adenoma, cholangiocarcinoma, or metastatic carcinomas to the liver. In human cancer of other anatomical sites, MXR7 mRNA was detected in low levels in one Wilms' tumor and in 4 of 40 gastric adenocarcinomas but not in several other types of cancer and 21 nonhepatocellular human tumor cell lines examined. MXR7 mRNA was expressed in high levels in the placenta, fetal liver, lung, and kidney, but it was undetectable in adult liver and was expressed in very low levels in adult lung and kidney. Our observations suggest that the MXR7 gene is regulated developmentally and expressed preferentially in HCC. To study its potential biological significance, we selected 113 patients who had unicentric primary HCC and had been followed for more than 4 years for further analysis. The MXR7 mRNA expression correlated closely with elevated serum alpha-fetoprotein (AFP) levels (88 versus 55%; P = 0.0001) and with expression of AFP mRNA (87 versus 55%; P = 0.005) and CD24 mRNA in HCC (80 versus 50%; P < 0.04), high tumor grade (76 versus 56%; P = 0.05), and tumor invasion (76 versus 55%; P < 0.05), but not with patient outcome. In HCC < or =3 cm, the frequency (77%) of MXR7 mRNA expression was significantly higher than that of elevated serum AFP (43%; P < 0.007) and AFP mRNA expression in HCC (41%; P < 0.004). Thus, MXR7 may serve as a sensitive early tumor marker for HCC and warrants more study to better understand its biological function.

Expression of p53 gene in 184 unifocal hepatocellular carcinomas: association with tumor growth and invasiveness.

To elucidate the biological significance of the p53 gene expression in human hepatocellular carcinoma (HCC), the p53 protein was studied in 184 resected unifocal primary HCCs, including 102 small (< or = 5 cm) and 82 large HCCs (> 5 cm), using immunocytochemistry. The p53 mRNA expression was analyzed in 69 cases using Northern hybridization. The p53 protein, which was detected in 58 HCCs (31.5%), was overexpressed more frequently in HCC with elevated serum alpha-fetoprotein level (37.9 versus 25%, P < 0.04), in large HCC (39.0 versus 25.5%, P < 0.03), and in invasive HCC (35.1 versus 13.3%, P < 0.01). The overexpression of p53 protein closely correlated with p53 mRNA overexpression (75 versus 44.4%, P < 0.003), and p53 gene mutation (76.9 versus 19.2%, P < 1 x 10(-9)). HCCs with p53 protein expression (group A) and those negative for both p53 protein and mRNA expression (group B) had an unfavorable outcome, while HCC with no p53 protein but with p53 mRNA overexpression (group C) had the best outcome; the 4-year survival was 26.1, 26.3, and 62.5%, respectively. The p53 gene mutation was significantly higher in group A HCC (76.9%) than in groups B (27.3%) and C (23.5%), P < 0.0001. The results suggest that the p53 protein and mRNA expression patterns in HCC correlate with p53 gene mutation and tumor behavior and may serve as a molecular prognostic factor.