Exploring the Effects of Probiotic Treatment on Urinary and Serum Metabolic Profiles in Healthy Individuals.

Probiotics are live microorganisms that confer health benefits when administered in adequate amounts. They are used to promote gut health and alleviate various disorders. Recently, there has been an increasing interest in the potential effects of probiotics on human physiology. In the presented study, the effects of probiotic treatment on the metabolic profiles of human urine and serum using a nuclear magnetic resonance (NMR)-based metabonomic approach were investigated. Twenty-one healthy volunteers were enrolled in the study, and they received two different dosages of probiotics for 8 weeks. During the study, urine and serum samples were collected from volunteers before and during probiotic supplementation. The results showed that probiotics had a significant impact on the urinary and serum metabolic profiles without altering their phenotypes. This study demonstrated the effects of probiotics in terms of variations of metabolite levels resulting also from the different probiotic posology. Overall, the results suggest that probiotic administration may affect both urine and serum metabolomes, although more research is needed to understand the mechanisms and clinical implications of these effects. NMR-based metabonomic analysis of biofluids is a powerful tool for monitoring host-gut microflora dynamic interaction as well as for assessing the individual response to probiotic treatment.

Probiotics as Potential Biological Immunomodulators in the Management of Oral Lichen Planus: What's New?

Oral lichen planus (OLP) is a T cell-mediated chronic inflammatory disorder with multifactorial aetiology and malignant transformation potential. Despite the treatments so far identified, new tailored and safe specific measures are needed. Recently, human microbiota imbalance has been linked to several immune-mediated diseases, opening new therapeutic perspectives for probiotics; besides their ability to directly interact with the host microbiota, they also display a strain-specific immune-modulatory effect. Thus, this non-systematic review aims to elucidate the molecular pathways underlying probiotic activity, mainly those of Lactobacilli and Bifidobacteria and their metabolites in OLP pathogenesis and malignant transformation, focusing on the most recent in vitro and in vivo research evidence. Findings related to their activity in other immune-mediated diseases are here included, suggesting a probiotic translational use in OLP. Probiotics show immune-modulatory and microbiota-balancing activities; they protect the host from pathogens, hamper an excessive effector T cell response, reduce nuclear factor-kappa B (NF-kB) signalling and basal keratinocytes abnormal apoptosis, shifting the mucosal response towards the production of anti-inflammatory cytokines, thus preventing uncontrolled damage. Therefore, probiotics could be a highly encouraging prevention and immunotherapeutic approach for a safer and more sustainable OLP management.

In Vitro Selection of Lactobacillus and Bifidobacterium Probiotic Strains for the Management of Oral Pathobiont Infections Associated to Systemic Diseases.

The human oral pathobionts Aggregatibacter actinomycetemcomitans, Streptococcus mitis and Streptococcus mutans, in dysbiosis-promoting conditions, lead to oral infections, which also represent a threat to human systemic health. This scenario may be worsened by antibiotic misuse, which favours multi-drug resistance, making the research on pathogen containment strategies more than crucial. Therefore, we aimed to in vitro select the most promising probiotic strains against oral pathogen growth, viability, biofilm formation, and co-aggregation capacity, employing both the viable probiotics and their cell-free supernatants (CFSs). Interestingly, we also assessed probiotic efficacy against the three-pathogen co-culture, mimicking an environment similar to that in vivo. Overall, the results showed that Lactobacillus CFSs performed better than the Bifidobacterium, highlighting Limosilactobacillus reuteri LRE11, Lacticaseibacillus rhamnosus LR04, Lacticaseibacillus casei LC04, and Limosilactobacillus fermentum LF26 as the most effective strains, opening the chance to deeper investigation of their action and CFS composition. Altogether, the methodologies presented in this study can be used for probiotic efficacy screenings, in order to better focus the research on a viable probiotic, or on its postbiotics, suitable in case of infections.

Oral microbiota and vitamin D impact on oropharyngeal squamous cell carcinogenesis: a narrative literature review.

An emerging body of research is revealing the microbiota pivotal involvement in determining the health or disease state of several human niches, and that of vitamin D also in extra-skeletal regions. Nevertheless, much of the oral microbiota and vitamin D reciprocal impact in oropharyngeal squamous cell carcinogenesis (OPSCC) is still mostly unknown. On this premise, starting from an in-depth scientific bibliographic analysis, this narrative literature review aims to show a detailed view of the state of the art on their contribution in the pathogenesis of this cancer type. Significant differences in the oral microbiota species quantity and quality have been detected in OPSCC-affected patients; in particular, mainly high-risk human papillomaviruses (HR-HPVs), Fusobacterium nucleatum, Porphyromonas gingivalis, Pseudomonas aeruginosa, and Candida spp. seem to be highly represented. Vitamin D prevents and fights infections promoted by the above identified pathogens, thus confirming its homeostatic function on the microbiota balance. However, its antimicrobial and antitumoral actions, well-described for the gut, have not been fully documented for the oropharynx yet. Deeper investigations of the mechanisms that link vitamin D levels, oral microbial diversity and inflammatory processes will lead to a better definition of OPSCC risk factors for the optimization of specific prevention and treatment strategies.

Non-Melanoma Skin Cancer: news from microbiota research.

Recently, research has been deeply focusing on the role of the microbiota in numerous diseases, either affecting the skin or other organs. What it is well established is that its dysregulation promotes several cutaneous disorders (i.e. psoriasis and atopic dermatitis). To date, little is known about its composition, mediators and role in the genesis, progression and response to therapy of Non-Melanoma Skin Cancer (NMSC). Starting from a bibliographic study, we classified the selected articles into four sections: i) normal skin microbiota; ii) in vitro study models; iii) microbiota and NMSC and iv) probiotics, antibiotics and NMSC. What has emerged is how skin microflora changes, mainly represented by increases of Staphylococcus aureus, Streptococcus pyogenes and Pseudomonas aeruginosa strains, modifications in the mutual quantity of ß-Human papillomavirus genotypes, of Epstein Barr Virus and Malassezia or candidiasis, may contribute to the induction of a state of chronic self-maintaining inflammation, leading to cancer. In this context, the role of S. aureus and that of specific antimicrobial peptides look to be prominent. Moreover, although antibiotics may contribute to carcinogenesis, due to their ability to influence the microbiota balance, specific probiotics, such as Lacticaseibacillus rhamnosus GG, Lactobacillus johnsonii NCC 533 and Bifidobacteria spp., may be protective.

Human papillomavirus type 16 E6 and E7 oncoproteins interact with the nuclear p53-binding protein 1 in an in vitro reconstructed 3D epithelium: new insights for the virus-induced DNA damage response.

BACKGROUND: Despite vaccination and screening measures, anogenital cancer, mainly promoted by HPV16 oncoproteins, still represents the fourth tumor and the second cause of death among women. Cell replication fidelity is the result of the host DNA damage response (DDR). Unlike many DNA viruses that promote their life cycle through the DDR inactivation, HR-HPVs encourage cells proliferation despite the DDR turned on. Why and how it occurs has been only partially elucidated. During HPV16 infection, E6 links and degrades p53 via the binding to the E6AP LXXLL sequence; unfortunately, E6 direct role in the DDR response has not clearly identified yet. Similarly, E7 increases DDR by competing with E2F1-pRb interaction, thus leading to the inactivation of pRb, and promotion, E2F1 mediated, of DDR genes translation, by binding to the pRb-like proteins CBP/p300 and p107, that also harbour LXXLL sequence, and via the interaction and activation of several DDR proteins. METHODS: To gain information regarding E6 and E7 contribution in DDR activation, we produced an in vitro 3D HPV16-E6E7 infected epithelium, already consolidated study model for HPVs, and validated it by assessing H&E staining and BrdU, HPV16 DNA, E6E7 proteins and γH2A.X/53BP1 double-strand break (DSBs) sensors expression; then we made an immuno-colocalization of E6 and E7 with cyclin E2 and B1. Since 53BP1, like E6 and E7, also binds p53 and pRb, we supposed their possible direct binding. To explore this hypothesis, we performed a double immunofluorescence of E6 and E7 with 53BP1, a sequence analysis of 53BP1 within its BRCT2 domain and then an in situ PLA within CaSki, E6E7HPV16 NHEKs and the 3D model. RESULTS: The in vitro epithelium resembled the histology and the events typical of in vivo infected tissues. E6E7HPV16 were both expressed in basal and differentiated strata and induced H2A.X phosphorylation and 53BP1 increment into nuclear foci. After highlighting E6 and E7 co-expression with 53BP1 and a LKVLL sequence within the 53BP1 BRCT2 domain, we demonstrated the bindings via the PLA technique. CONCLUSIONS: Our results reinforce E6 and E7 role in cellular function control providing potentially new insights into the activity of this tumor virus.

Rutin, a Quercetin Glycoside, Restores Chemosensitivity in Human Breast Cancer Cells.

Several studies have documented the ability of flavonoids to sensitize cancer cells to chemotherapeutics and reverse multidrug resistance by inhibition of efflux pumps (adenosine triphosphate-binding cassette transporters), apoptosis activation, and cell cycle arrest. In this study, the flavonoid rutin (quercetin 3-O-ß-d-rutinoside) was investigated as chemosensitizer towards two different human epithelial breast cancer cell lines: (i) MB-MDA-231, selected as representative for triple-negative breast cancer and (ii) MCF-7 used as a well-characterized model of HER2-negative breast cancer. To assess the cytocompatibility of rutin against non-cancer cells, primary human mammary fibroblasts were used as control and non-target cells. In MDA-MB-231 cells, 20 µM rutin enhanced cytotoxicity related to cyclophosphamide and methotrexate. Rutin significantly (p < 0.05) increased the anticancer activity of both chemotherapeutics, at 24-48-72 h, and decreased the activity of the adenosine triphosphate-binding cassette transporters, namely, P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP). Flow cytometry analysis showed 20 µM and 50 µM rutin arrested cell cycle at G2/M and G0/G1 phases, respectively, significantly promoting cell apoptosis. Rutin, via non-selective inhibition of P-gp and BCRP pumps, efficiently reverses multidrug resistance and restores chemosensitivity to cyclophosphamide and cyclophosphamide of human chemoresistant, triple-negative breast cancer cells, successfully arresting cell cycle progression. Copyright © 2017 John Wiley & Sons, Ltd.

Essential Oil from Berries of Lebanese Juniperus excelsa M. Bieb Displays Similar Antibacterial Activity to Chlorhexidine but Higher Cytocompatibility with Human Oral Primary Cells.

Chlorhexidine (CHX), one of the most effective drugs administered for periodontal treatment, presents collateral effects including toxicity when used for prolonged periods; here, we have evaluated the bactericidal potency and the cytocompatibility of Juniperus excelsa M. Bieb essential oil (EO) in comparison with 0.05% CHX. The EO was extracted from berries by hydrodistillation and components identified by gas chromatography and mass spectrometry. Bacterial inhibition halo analysis, quantitative cell viability 2,3-bis(2-methoxy-4-nitro-5-sulphophenyl)-5-[(phenyl amino) carbonyl]-2H-tetrazolium hydroxide assay (XTT), and colony forming unit (CFU) count were evaluated against the two biofilm formers Aggregatibacter actinomycetemcomitans and Streptococcus mutans. Finally, cytocompatibility was assessed with human primary gingival fibroblasts (HGF) and mucosal keratinocytes (HK). The resulting EO was mainly composed of monoterpene hydrocarbons and oxygenated monoterpenes. An inhibition halo test demonstrated that both bacteria were sensitive to the EO; XTT analysis and CFU counts confirmed that 10-fold-diluted EO determined a statistically significant (p < 0.05) reduction in bacteria count and viability towards both biofilm and planktonic forms in a comparable manner to those obtained with CHX. Moreover, EO displayed higher cytocompatibility than CHX (p < 0.05). In conclusion, EO exhibited bactericidal activity similar to CHX, but a superior cytocompatibility, making it a promising antiseptic alternative to CHX.

Adipose-derived adult stem cells: available technologies for potential clinical regenerative applications in dentistry.

Tissue homeostasis depends closely on the activity and welfare of adult stem cells. These cells represent a promising tool for biomedical research since they can aid in treatment and promote the regeneration of damaged organs in many human disorders. Adult stem cells indefinitely preserve their ability to self-renew and differentiate into various phenotypes; this capacity could be promoted in vitro by particular culture conditions (differentiation media) or spontaneously induced in vivo by exploiting the biochemical and mechanical properties of the tissue in which the stem cells are implanted. Among the different sources of adult stem cells, adipose tissue is an attractive possibility thanks to its ready availability and the standard extraction techniques at our disposal today. This review discusses the isolation, characterization, and differentiation of human adipose-derived adult stem cells, as well as regeneration strategies, therapeutic uses, and adverse effects of their delivery. In particular, since oral disorders (e.g., trauma, erosion, and chronic periodontitis) often cause the loss of dental tissue along with functional, phonetic, and aesthetic impairment, this review focuses on the application of human adipose-derived adult stem cells, alone or in combination with biomaterials, in treating oral diseases.

Tissue-engineered skin substitutes: an overview.

Extensive skin loss and chronic wounds are still a significant challenge to clinicians: even if injured epidermis is normally able to self-renew, deep injuries can cause negative regulation of the wound healing cascade, leading to chronic wound formation. Skin-autografting surgical procedures are often limited by the poor availability of healthy tissue, whereas the use of non-self-tissues for allografts presents some severe risks. Tissue-engineered skin substitutes have recently become viable as a suitable alternative to auto- and allografts. However, biologists, biochemists, and technical engineers are still struggling to produce complex skin substitutes that can readily be transplanted in large quantities. The ambitious goal is now to construct a dermoepidermal substitute that rapidly vascularizes and optimally supports a stratifying epidermal graft on a biodegradable matrix. This review analyzes these aspects in light of the available literature and the authors' experience.

Role of the Microbiota in Skin Neoplasms: New Therapeutic Horizons.

The skin and the gut are regularly colonized by a variety of microorganisms capable of interacting with the immune system through their metabolites and influencing the balance between immune tolerance and inflammation. Alterations in the composition and diversity of the skin microbiota have been described in various cutaneous diseases, including skin cancer, and the actual function of the human microbiota in skin carcinogenesis, such as in progression and metastasis, is currently an active area of research. The role of Human Papilloma Virus (HPV) in the pathogenesis of squamous cell carcinoma is well consolidated, especially in chronically immunosuppressed patients. Furthermore, an imbalance between Staphylococcus spp., such as Staphylococcus epidermidis and aureus, has been found to be strongly related to the progression from actinic keratosis to squamous cell carcinoma and differently associated with various stages of the diseases in cutaneous T-cell lymphoma patients. Also, in melanoma patients, differences in microbiota have been related to dissimilar disease course and prognosis and may affect the effectiveness and tolerability of immune checkpoint inhibitors, which currently represent one of the best chances of a cure. From this point of view, acting on microbiota can be considered a possible therapeutic option for patients with advanced skin cancers, even if several issues are still open.

Growth Conditions Influence Lactobacillus Cell-Free Supernatant Impact on Viability, Biofilm Formation, and Co-Aggregation of the Oral Periodontopathogens Fusobacterium nucleatum and Porphyromonas gingivalis.

Fusobacterium nucleatum and Porphyromonas gingivalis human periodontopathogens play a leading part in oral squamous cell carcinoma through cell proliferation, invasion, and persistent inflammation promotion and maintenance. To explore how the activity of Lactobacillus-derived cell-free supernatants (CFSs) can be influenced by growth medium components, CFSs were produced both in the standard MRS and the novel animal-derivative-free "Terreno Industriale Lattobacilli" (TIL) media, and in vitro screened for the containment of F. nucleatum and P. gingivalis both single and co-cultured and also for the interference on their co-aggregation. The viability assay demonstrated that the Limosilactobacillus reuteri LRE11 and Ligilactobacillus salivarius LS03 MRS-produced CFSs were significantly more effective against single and co-cultured pathogens. All the other CFSs significantly improved their efficacy when produced in TIL. Both MRS- and TIL-produced CFSs significantly inhibited the single and co-cultured pathogen biofilm formation. Only Levilactobacillus brevis LBR01 CFS in MRS specifically reduced F. nucleatum and P. gingivalis co-aggregation, while viable LBR01, LS03, and LRE11 in MRS significantly co-aggregated with the pathogens, but only LS03 cultivated in TIL improved this effect. This work paves the way to better consider environmental growth conditions when screening for probiotic and postbiotic efficacy as crucial to pathogen aggregation, adhesion to the host's niches, and exclusion.

Lactobacillus johnsonii LJO02 (DSM 33828) Cell-Free Supernatant and Vitamin D Improve Wound Healing and Reduce Interleukin-6 Production in Staphylococcus aureus-Infected Human Keratinocytes.

Methicillin-resistant biofilm-forming Staphylococcus spp. are found in about 25% of the overall cases of chronic wounds, which can undergo malignant degeneration and be associated with skin cancer. Although antimicrobial agents are clinically used to counteract pathogens and promote wound healing, they are increasingly ineffective against multi-drug resistant bacteria. Moreover, they can induce dysbiosis, which favors opportunistic pathogen infections and alters immune responses. Consequently, research on pathogen containment strategies is crucial. We aimed to evaluate the potential beneficial effect of Lactobacillus johnsonii LJO02 cell-free supernatant (CFS) and vitamin D, as single treatments or in combination, on cell viability, wound healing, and the pro-inflammatory interleukin-6 (IL-6) production of a Staphylococcus aureus-infected human immortalized keratinocyte cell line (HaCaT) in vitro model. The analysis showed that LJO02 CFS 20% v/v ratio and 100 nM vitamin D promoted infected cell viability and wound healing and significantly reduced IL-6 production. However, their effect was not synergic, since no significant difference between the single and combined treatments was observed. LJO02 CFS topic application and vitamin D supplementation could provide a valuable strategy for attenuating S. aureus-induced pathogenesis, promoting wound healing and opening new therapeutic strategies supporting the conventional approaches.

Microbiota, Oxidative Stress, and Skin Cancer: An Unexpected Triangle.

Mounting evidence indicates that the microbiota, the unique combination of micro-organisms residing in a specific environment, plays an essential role in the development of a wide range of human diseases, including skin cancer. Moreover, a persistent imbalance of microbial community, named dysbiosis, can also be associated with oxidative stress, a well-known emerging force involved in the pathogenesis of several human diseases, including cutaneous malignancies. Although their interplay has been somewhat suggested, the connection between microbiota, oxidative stress, and skin cancer is a largely unexplored field. In the present review, we discuss the current knowledge on these topics, suggesting potential therapeutic strategies.

Distinct Signatures of Tumor-Associated Microbiota and Metabolome in Low-Grade vs. High-Grade Dysplastic Colon Polyps: Inference of Their Role in Tumor Initiation and Progression.

According to the driver-passenger model for colorectal cancer (CRC), the tumor-associated microbiota is a dynamic ecosystem of bacterial species where bacteria with carcinogenic features linked to CRC initiation are defined as "drivers", while opportunistic bacteria colonizing more advanced tumor stages are known as "passengers". We reasoned that also gut microbiota-associated metabolites may be differentially enriched according to tumor stage, and be potential determinants of CRC development. Thus, we characterized the mucosa- and lumen-associated microbiota (MAM and LAM, respectively) and mucosa-associated metabolites in low- vs. high-grade dysplastic colon polyps from 78 patients. We show that MAM, obtained with a new biopsy-preserving approach, and LAM differ in composition and α/ß-diversity. By stratifying patients for polyp histology, we found that bacteria proposed as passengers by previous studies colonized high-grade dysplastic adenomas, whereas driver taxa were enriched in low-grade polyps. Furthermore, we report altered "mucosa-associated metabolite" levels in low- vs. high-grade groups. Integrated microbiota-metabolome analysis suggests the involvement of the gut microbiota in the production and consumption of these metabolites. Altogether, our findings support the involvement of bacterial species and associated metabolites in CRC mucosal homeostasis in a tumor-stage-specific manner. These distinct signatures may be used to distinguish low-grade from high-grade dysplastic polyps.

An animal derivative-free medium enhances Lactobacillus johnsonii LJO02 supernatant selective efficacy against the methicillin (oxacillin)-resistant Staphylococcus aureus virulence through key-metabolites.

The spread of multidrug-resistant bacteria, such as the skin commensal Staphylococcus aureus, is a worldwide health challenge; new methods to counteract opportunistic pathogen growth and virulence are urgent. We compared the activity of Lacticaseibacillus rhamnosus LR06 (DSM 21981) and Lactobacillus johnsonii LJO02 (DSM 33828) cell-free supernatants (CFSs) produced in the conventional animal derivative-based MRS medium and an innovative animal derivative-free broth (TIL) versus the MDR S. aureus (ATCC 43300). CFS influence was assessed towards the viability, metabolic activity, and ability to form biofilm of the MDR strain through optical density, alamarBlue assay, and crystal violet staining; their content in short-chain fatty acids, lactic acid, and proteins was analysed via high-resolution mass spectrometry and gas chromatography. All CFSs reduce viable and metabolically active S. aureus, being TIL more efficient compared to MRS in stimulating lactic acid bacteria metabolism and decreasing S. aureus biofilm formation. Particularly, the CFS from LJO02 grown in TIL has the best efficacy, revealing a high amount of lactic acid and 59 peculiar proteins; its effectiveness is partially maintained upon trypsin and proteinase K treatments, but not by pepsin and pH basification. Therefore, antagonistic CFSs may represent a strategic prevention approach, with bacteriotherapeutic and bio-repair potential.

Macrophages expressing TREM-1 are involved in the progression of HPV16-related oropharyngeal squamous cell carcinoma.

INTRODUCTION: Many types of research have been performed to improve the diagnosis, therapy, and prognosis of oropharyngeal carcinomas (OP-SCCs). Since they arise in lymphoid-rich areas and intense lymphocytic infiltration has been related to a better prognosis, a TREM-1 putative function in tumour progression and survival has been hypothesized. MATERIALS AND METHODS: Twenty-seven human papillomavirus (HPV) 16+ OP-SCC specimens have been analyzed to relate TREM-1 expression with histiocytic and lymphocytic markers, HPV presence and patients' outcome. RESULTS: No differences have been shown between intratumoral and stromal CD4+ cells, while intratumoral CD8+ lymphocytes are higher with respect to the tumour stroma (p = .0005). CD68+ cells are more than CD35+ and TREM-1+; their presence is related to CD35± and TREM-1± histiocytes (p = .005 and .026, respectively). Intratumoral CD4+ lymphocytes are higher in p16+ cases (11/27) than in p16- (p = .042); moreover, p16 positivity correlates to a better survival (p = .034). CD4+, CD8+ and CD35+ cells have no impact on survival, while CD68 expression heavily influences progression and bad outcome (p = .037). TREM-1 positivity also leads to worst overall survival (p = .001): peritumoral expression and death-cause relationship are always significant, particularly when the cause is OP-SCC (p = .000). CONCLUSION: While p16 shows to better stratify HPV16+ patients' outcome, TREM-1+ macrophages suggest their key importance in HPV-related OP-SCCs progression.KEY MESSAGESTREM-1 positivity correlates to the worst overall survival of HPV16-positive OPSCCs-affected patients.p16-positive HPV16 related OPSCCs patients have a better prognosis with respect to p16-negative ones.

Effects of Probiotics Administration on Human Metabolic Phenotype.

The establishment of the beneficial interactions between the host and its microbiota is essential for the correct functioning of the organism, since microflora alterations can lead to many diseases. Probiotics improve balanced microbial communities, exerting substantial health-promoting effects. Here we monitored the molecular outcomes, obtained by gut microflora modulation through probiotic treatment, on human urine and serum metabolic profiles, with a metabolomic approach. Twenty-two subjects were enrolled in the study and administered with two different probiotic types, both singularly and in combination, for 8 weeks. Urine and serum samples were collected before and during the supplementation and were analyzed by nuclear magnetic resonance (NMR) spectroscopy and statistical analyses. After eight weeks of treatment, probiotics deeply influence the urinary metabolic profiles of the volunteers, without significantly altering their single phenotypes. Anyway, bacteria supplementation tends to reduce the differences in metabolic phenotypes among individuals. Overall, the effects are recipient-dependent, and in some individuals, robust effects are already well visible after four weeks. Modifications in metabolite levels, attributable to each type of probiotic administration, were also monitored. Metabolomic analysis of biofluids turns out to be a powerful technique to monitor the dynamic interactions between the microflora and the host, and the individual response to probiotic assumption.

Expression of the interferon-inducible proteins MxA and IFI16 in liver allografts.

AIMS: To test the hypothesis that the activation of the interferon (IFN) system pathways might link hepatitis C virus (HCV) recurrence in the liver allograft with acute cellular rejection. METHODS AND RESULTS: In this retrospective study, allograft biopsy specimens from 28 adult patients (14 HCV+ and 14 HCV-) who had undergone their first liver transplantation were analysed. Eleven biopsy specimens showed acute cellular rejection (Banff rejection activity index score > or =3). Specimens were immunostained for two IFN-inducible proteins, MxA and IFI16, and for CD45. The predominant MxA reactivity pattern was hepatocytic, whereas IFI16 was expressed in both the hepatocellular and inflammatory compartments. Moderate to strong MxA expression in hepatocytes was associated positively with rejection score (P < 0.01), donor's age < or =45 years (P < 0.05) and aspartate aminotransferase levels >40 U/l on the day of biopsy (P < 0.05), and inversely with infiltration of portal triads by IFI16+/CD45+ cells (P < 0.005) and time to progression beyond Ishak stage 2 of recurrent hepatitis C (P < 0.01). On multivariate analysis, MxA expression in hepatocytes was independently associated with allograft rejection and donor's age. CONCLUSIONS: Acute allograft rejection and recurrence of HCV infection in the liver allograft appear to intersect in the IFN system pathways.

Proteomic investigation in A549 lung cell line stably infected by HPV16E6/E7 oncogenes.

BACKGROUND: Data have accumulated implicating the involvement of oncogenic human papillomaviruses (HPVs) in bronchial carcinogenesis. We recently described the presence of oncogenic HPV transcripts in non-small cell lung cancers. OBJECTIVE: To investigate the role of oncogenic HPVs in lung carcinogenesis. MATERIAL AND METHODS: The lung cell line A549 stably infected with HPV16E6, HPV16E7 and HPVE6/E7 constructs was used to investigate the protein profile changes associated with the expression of these oncogenes. Replicated two-dimensional gel electrophoresis gels from uninfected and stably HPV16E6-, E7-, and E6/E7-infected A549 cells were compared for changes in protein profile. Protein identification was achieved by peptide mass fingerprinting by MALDI-TOF-MS and nLC-ESI-Q-TOF-MS/MS peptide ladder sequencing. RESULTS: We identified 17 different polypeptides whose average normalized spot intensity was statistically significant (p < 0.05) and differed by 2-fold. Relationships between differentially expressed proteins and the HPV-induced infection mechanism have been clustered by knowledge-base database functional association network analysis. CONCLUSION: The impact of Hsp27, annexin III, annexin IV, Gp96 and TPT1 on the cellular response mechanism to HPV infection is presented and discussed.