[A prospective cohort study on the effect of implant restoration following micro crestal flap-alveolar ridge preservation at molar extraction sockets with severe periodontitis].

Objective: To evaluate the survival rate, success rate, soft tissue conditions and marginal bone level changes of implants following micro crestal flap-alveolar ridge preservation at molar extraction sockets with severe periodontitis, compared to natural healing. Methods: From March 2015 to January 2017, patients scheduled for molar extraction as a consequence of severe periodontitis and planned implant-retained prostheses from Department of Periodontology Peking University School and Hospital of Stomatology were selected. A total of 40 molar extraction sockets from 40 patients received implant placement following micro crestal flap-alveolar ridge preservation or natural healing. The front consecutive 20 teeth were assigned to the natural healing group, and the back ones were assigned to the micro crestal flap-alveolar ridge preservation (MCF-ARP) group. The superstructures were placed 6 months later. Within 2 weeks (baseline) and 1, 2 and 3 years after implant crown restoration, modified plaque index, probing depth, modified bleeding index and keratinized tissue width were recorded every six months. Parallel periapical radiographs were taken to evaluate the peri-implant marginal bone level and to calculate marginal bone loss. Independent sample t test or Mann-Whitney U test were used to compare the differences in the above clinical and imaging indicators between the two groups. Results: The implant survival rate and success rate of the two groups were both 100% (20/20). There were no significant differences in the modified plaque index, probing depth, modified bleeding index, buccal keratinized tissue width and marginal bone loss between two groups at 1, 2 and 3 years after implant crown restoration (all P>0.05). Marginal bone loss was 0.22 (0.14, 0.34) mm in the natural healing group and 0.21 (0.12, 0.30) mm in the MCF-ARP group at a 3-year post-loading evaluation. Conclusions: Within the limitations of the present study, implants placed at ridge preserved and naturally healed molar extraction sockets with severe periodontitis demonstrate comparable clinical outcomes at a 3-year post-loading evaluation.

[Profiles and differences of submucosal microbial in peri-implantitis and health implants: A cross-sectional study].

OBJECTIVE: To describe the submucosal microbial profiles of peri-implantitis and healthy implants, and to explore bacteria that might be correlated with clinical parameters. METHODS: In the present cross-sectional study, 49 patients were recruited. Each patient contributed with one implant, submucosal biofilms were collected from 20 healthy implants and 29 implants with peri-implantitis. DNA was extracted and bacterial 16S ribosomal RNA (16S rRNA) genes were amplified. Submucosal biofilms were analyzed using 16S rRNA sequencing at Illumina MiSeq platform. Differences between the groups were determined by analyzing α diversity, microbial component and microbial structure. The potential correlation between the bacteria with pocket probing depth (PPD) of peri-implant calculated by Spearman correlation analysis. RESULTS: The α diversity of submucosal microbial of health group was significantly lower than that in peri-implantitis group (Chao1 index: 236.85±66.13 vs. 150.54±57.43, P < 0.001; Shannon index: 3.42±0.48 vs. 3.02±0.65, P=0.032). Principal coordinated analysis showed that the submucosal microbial structure had significant difference between healthy and peri-implantitis groups [R2=0.243, P=0.001, analysis of similarities (ANOSIM)]. Compared with healthy implants, relative abundance of periodontal pathogens were higher in peri-implantitis, including members of the red complex (Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola) and some members of orange complex (Precotella intermedia, Eubacterium nodatum, Parvimonas micra), as well as some new periodontal pathogens, such as Fillifactor alocis, Fretibacterium fastidiosum, Desulfobulbus sp._HMT_041, and Porphyromonas endodontalis. Spearman correlation analysis revealed that the relative abundance of Treponema denticola (r=0.686, P < 0.001), Tannerella forsythia (r=0.675, P < 0.001), Fretibacterium sp. (r=0.671, P < 0.001), Desulfobulbus sp._HMT_041 (r=0.664, P < 0.001), Filifactor alocis (r=0.642, P < 0.001), Fretibacterium fastidiosum (r=0.604, P < 0.001), Porphyromonas gingivalis (r=0.597, P < 0.001), Porphyromonas endodontalis (r=0.573, P < 0.001) were positive correlated with PPD. While the relative abundance of Rothia aeria (r=-0.615, P < 0.001) showed negatively correlation with PPD. CONCLUSION: Marked differences were observed in the microbial profiles of healthy implants and peri-implantitis. The members of red and orange complex as well as some new periodontal pathogens seem to play an important role in peri-implant disease. Compared with healthy implants, the submucosal microbial of peri-implantitis were characterized by high species richness and diversity.

[Healing of the dento-gingival junction following modified crown lengthening procedure in beagle dogs].

OBJECTIVE: To evaluate the type of wound healing following modified crown lengthening surgery in dog model to provide a biological basis for its clinical application. METHODS: Flap surgery, traditional crown lengthening procedure and modified crown lengthening procedure were performed on the right maxillary central incisor, the left maxillary central incisor and the left maxillary first lateral incisor respectively of five male beagle dogs. The right maxillary first lateral incisors with no surgical intervention were used as controls. Thirty-six weeks after the experimental procedure, tissue blocks were harvested and prepared for histological examination and analysis. RESULTS: Histometric examination of buccolingual sections stained with hematoxylin-eosin demonstrated that the type of wound healing in the flap surgery group was re-attachment, similar to the control group. For the traditional crown lengthening surgery group, all of the five beagle dogs had lamellar cementum defects on root surface, the wound healing of four beagle dogs was new attachment accompanied by new cementum formation at cementum defect areas and the suprac-restal connective tissue was functionally oriented perpendicular to the new cementum. The wound healing of the other beagle dog was long junctional epithelial attachment, in which the junctional epithelium extended to the apical terminus of the cementum defect. In the modified crown lengthening surgery group, four beagle dogs had cementum defects on root surface (two lamellar cementum defects and two shallow platform-like cementum defects), the wound healing of three beagle dogs was new attachment, however, the supracrestal connective tissue was parallel to the root surface. The type of wound healing of another one beagle dog was long junctional epithelial attachment. Wound healing of one beagle dog in this group could not be characterized due to incomplete dissection. CONCLUSION: Wound healing in the modified crown lengthening surgery group was similar to the traditional crown lengthening surgery group, and two types of wound healing were observed: new attachment and long junctional epithelium attachment. Neither type of root treatment procedure (root planing or root reshaping) nor root surface defect pattern (the lamellar cementum defect or shallow platform-like cementum defect) influenced the observed type of wound healing.

[Analysis the influence factors of treatment free remission outcome with chronic myeloid leukemia patients who discontinued tyrosine kinase inhibitors].

Objective: To explore the related factors affecting the outcome of treatment free remission (TFR) in patients with chronic myeloid leukemia (CML). Methods: Clinical data of CML patients with automatic discontinuation of tyrosine kinase inhibitor (TKI) from the CML cooperative organization of Henan province between June 2, 2013 to March 27, 2021 and the follow-up time was ≥ 6 months were retrospectively analyzed. Log-rank test was used for univariate analysis and Cox proportional risk regression model was used for multivariate analysis. Results: A total of 135 patients were enrolled, and 69 patients (51.1%) were femal and 66 patients (48.9%)were male. Median age was[M(Q1,Q3)] 49 years (38, 58)at discontinuation.Before discontinuation, 72 patients (53.3%) were on treatment with second-generation TKI, 63 patients (46.7%) were on treatment with IM, 17patients (12.6%) had a history of TKI reduction/withdrawal;median duration of treatment was months 84 (68, 108) for all patients;median time of TKI treatment to DMR was months 12(8, 26);median duration of DMR was months 65 (54, 84), and 9 patients (6.7%) had unsustained DMR.The median follow-up time was months 16(6-96), 35 patients (25.9%) lost MMR at a median months 3(1-22), overall estimated TFR was 74.1%.The univariate analysis results showed that:second-generation TKI was used, the time of TKI treatment to DMR was ≤12 months, DMR duration time ≥48 months, had sustained DMR, without TKI reduction/withdrawal history were favorable factors affecting of TFR in patients with TKI discontinuation (all P<0.05).The TFR rate of the second-generation TKI therapy group was significantly higher than the IM therapy group (81.9% vs 65.1%, P=0.019).The multivariate analysis results showed that second-generation TKI treatment[RR=0.451, 95%CI (0.227-0.896), P=0.023] and had sustained DMR [RR=0.120, 95%CI (0.053-0.271), P<0.001] were the protective factors of TFR in patients with TKI discontinuation. Conclusions: Treated with second-generation TKI and had sustained DMR are the protective factors of TFR in patients with TKI discontinuation.The CML patients who had sustained DMR more≥48 months before TKI discontinuation showed a better TFR.

[Efficacy of combined application of glycine powder air-polishing in non-surgical treatment of peri-implant diseases].

OBJECTIVE: To compare the clinical efficacy of combined application of glycine powder air-polishing and mechanical submucosal debridement in non-surgical treatment of peri-implant diseases. METHODS: A randomized controlled clinical study was carried out on patients diagnosed with peri-implant diseases in the Department of Periodontology, Peking University School and Hospital of Stomatology, between May of 2020 and June of 2021.Twenty-eight patients with totally sixty-two implants were enrolled.The patients were randomly divided into the test group and control group. The patients in the test group (13 subjects/32 implants) received mechanical submucosal debridement using titanium curettes combined with application of glycine powder air-polishing, while the control group (15 subjects/30 implants) received mechanical submucosal debridement using titanium only. Clinical parameters, such as plaque index (PLI), pocket probing depth (PPD), bleeding index (BI) and the percentage of suppuration on probing on implants' level (SoP%) were measured at baseline and 8 weeks after non-surgical intervention. Changes and group differences of clinical parameters of the implants before and 8 weeks after non-surgical intervention were compared. RESULTS: Mean PLI, PPD, BI of both the test group and control group significantly reduced 8 weeks after non-surgical intervention (P < 0.05). Compared with the control group, the test group achieved lower BI (2.7±0.8 vs. 2.2±0.7, P < 0.05), more reduction of BI (0.6±0.7 vs. 1.1±0.6, P < 0.01) and more reduction of SoP% (21.9% vs. 10%, P < 0.05) after non-surgical intervention. Both the control and test groups exhibited comparable PLI and PPD reductions (P>0.05). For the implants diagnosed with peri-implant mucositis, the test group revealed more signi-ficant reduction in BI and SoP% than the control group (1.0±0.7 vs. 0.4±0.7, P=0.02; 6.3% vs. 0, P=0.012). There was no significant difference existing in PLI and PD improvement between the control group and test group (P>0.05). For the implants diagnosed with peri-implantitis, there was no significant difference existing in PLI, PPD, BI and SoP% improvement values between the test and control groups (P>0.05). No complications or discomforts were reported during the study. CONCLUSION: Both treatment procedures could relieve the inflammation of peri-implant soft tissue. Non-surgical mechanical submucosal debridement combined application of glycine powder air-polishing is associated with significant reduction of soft tissue bleeding and suppuration on probing especially in the implants diagnosed with peri-implant mucositis.

[Targeted immunotherapy efficacy analysis in patients with relapsed/refractory B cell acute lymphocytic leukemia].

Objective: Comparison of conventional chemotherapy and immunotargeted therapy efficacy in patients with relapsed/refractory (R/R) acute B cell leukemia (B-ALL) . Methods: The clinical data of 212 patients with R/R B-ALL in the Affiliaed Cancer Hospital of Zhengzhou University from January 2008 to July 2020 were analyzed retrospectively to compare the response rate and survival time difference between conventional chemotherapy and immunotargeted therapies (antiCD19 CAR-T and CD3CD19 bi-specific antibody blinatumomab) , and to explore the related factors affecting prognosis. Results: The CR rate of patients with R/R B-ALL treated with anti-CD19 CAR-T cells was 80.4% , patients treated with blinatumomab was 62.5% , and patients treated with chemotherapy was 38.6% . There was significant difference in the CR rate among the three therapies (P<0.001) . CAR-T cells 1-year OS rate was 41.5% , which was significantly higher than that of the chemotherapy group (10.3% ) (P<0.001) . The 1-year PFS rate of CAR-T cells (30.1% ) was also significantly higher than that of the chemotherapy group (9.7% ) (P<0.001) . The median OS of patients with bridging allo-HSCT after CR treatment by CAR-T cells was 18.5 months, which was higher than that of patients without allo-HSCT (8 months) (P=0.027) . The median PFS of patients with allo-HSCT was 17 months, which was higher than that of patients without allo-HSCT (4 months) (P=0.001) . The 1-year OS rate of patients treated with blinatumomab was 14.3% , which was higher than that of the chemotherapy group (10.3% ) (P=0.018) . The 1-year PFS rate (14.6% ) was also higher than that of the chemotherapy group (9.7% ) (P=0.046) . The median OS and median PFS of patients with bridging allo-HSCT were 13 and 11 months, respectively, which was higher than that of patients without allo-HSCT (9.5 and 6 months) . The cytokine release syndrome (CRS) incidence in patients with R/R B-ALL treated with anti-CAR-T cells was 89.8% . Grades 3-4, grade 2, and grade 1 CRS were experienced by 30.2% , 11.3% and 58.5% patients, respectively. Only three patients (37.5% ) with blinatumomab developed CRS, all of which were grade 1. Conclusion: The response rate and survival rate of patients with R/R B-ALL treated with CD19 CAR-T cells and blinatumomab were significantly better than those treated with conventional chemotherapy.

[Two procedures for ridge preservation of molar extraction sites affected by severe bone defect due to advanced periodontitis].

OBJECTIVE: To evaluate and to compare dimensional alterations of hard and soft tissues in molar extraction sites with irregular deficiency of bone plates due to advanced periodontitis receiving two different procedures, namely the flapped and flapless techniques with Bio-Gide membrane covering the Bio-Oss material for ridge preservation. METHODS: Twenty-three patients with 24 infected-molar extraction sites received ridge preservation procedure, the first consecutive 12 sites belonged to the flap group (a full thickness mucoperiosteal flap and primary soft tissue closure) and the following 12 sites belonged to the flapless group (minimal flap with a collagen sponge and a secondary soft tissue closure). Width of keratinized tissue was evaluated before tooth extraction and after 6-month healing. Parallel periapical radiographs were taken immediately and 6 months after extraction to evaluate vertical bone changes. The width of the ridge was measured in the center of the ridge at the time of tooth extraction and after 6 months at implant placement. RESULTS: After 6 months, width of keratinized tissue decreased (1.6±1.5) mm in the flap group (P=0.004) when compared with (0.3±1.6) mm in the flapless group (P>0.05). Both groups showed increases in ridge height from the central aspect, (5.53±4.20) mm for flap group and (7.70±4.35) mm for flapless group. These differences between the groups were not statistically significant (P=0.226). The ridge widths were (9.5±2.2) mm for flap group and (9.3±1.0) mm for flapless group at the time of implant insertion, and no statistical significance was observed between the flap and flapless groups. CONCLUSION: The study points out that both ridge preservation techniques were effective in increasing ridge height and minimizing ridge resorption after tooth extraction, and the ridge width allowed the placement of implants 6 months after ridge preservation. The flapless technique gave positive outcome in terms of the keratinized gingival width than that of the flap technique.

Periodontal evaluation of crown-root fractured teeth following modified crown lengthening surgery.

Objectives To evaluate long-term outcomes of modified crown lengthening procedures for crown-root fractured teeth, and to analyse factors that affect the periodontal health of the treated teeth.Methods The present study retrospectively analysed data from 22 patients (13 males and 9 females) who had presented with crown-root fractured teeth and received modified crown lengthening procedures combined with crown restoration for ≥13 months. In total, 26 crown-root fractured teeth were treated and examined for plaque index, probing depth, bleeding index, bleeding on probing, mobility and the crown margin location. The contralateral tooth and ipsilateral teeth mesial and distal to the treated tooth were used as controls. Data were assessed descriptively or analysed statistically with Mann-Whitney-U test at α=0.05.Results Recorded periodontal indices revealed stable periodontal status in 25 of 26 treated teeth with the mean values for aesthetic and functional VAS scores at 9.5. A negative correlation was observed between the subgingival crown margin location and the bleeding index.Conclusions The modified crown lengthening procedure is a feasible and minimally invasive therapeutic option for management of crown-root fractured cases.

[Dental implantation and soft tissue augmentation after ridge preservation in a molar site: a case report].

For ideal implant rehabilitation, an adequate bone volume, optical implant position, and stable and healthy soft tissue are required. The reduction of alveolar bone and changes in its morphology subsequent to tooth extraction will result in insufficient amount of bone and adversely affect the ability to optimally place dental implants in edentulous sites. Preservation of alveolar bone volume through ridge preservation has been demonstrated to reduce the vertical and horizontal contraction of the alveolar bone crest after tooth extraction and reduce the need for additional bone augmentation procedures during implant placement. In this case, a patient presented with a mandible molar of severe periodontal disease, the tooth was removed as atraumatically as possible and the graft material of Bio-Oss was loosely placed in the alveolar socket without condensation and covered with Bio-Gide to reconstruct the defects of the alveolar ridge. Six months later, there were sufficient height and width of the alveolar ridge for the dental implant, avoiding the need of additional bone augmentation and reducing the complexity and unpredictability of the implant surgery. Soft tissue defects, such as gingival and connective tissue, played crucial roles in long-term implant success. Peri-implant plastic surgery facilitated development of healthy peri-implant structure able to withstand occlusal forces and mucogingival stress. Six months after the implant surgery, the keratinized gingiva was absent in the buccal of the implant and the vestibular groove was a little shallow. The free gingival graft technique was used to solve the vestibulum oris groove supersulcus and the absence of keratinized gingiva around the implant. The deepening of vestibular groove and broadening of keratinized gingiva were conducive to the long-term health and stability of the tissue surrounding the implant. Implant installation and prosthetic restoration showed favorable outcome after six months.

[Finite element analysis of the maxillary central incisor with traditional and modified crown lengthening surgery and post-core restoration in management of crown-root fracture].

OBJECTIVE: To construct three-dimensional finite element models with modified crown lengthening surgery and post-core restoration in management of various crown-root fracture types, to investigate the intensity and distribution of stressin models mentioned above, and to compare and analyze the indications of traditional and modified crown lengthening surgeries from the mechanic point of view. METHODS: Nine three-dimensional finite element models with modified crown lengthening surgery and post-core restoration were established and analyzed by micro-CT scanning technique, dental impression scanner, Mimics 10.0, Geomagic studio 9.0 and ANSYS 14.0 software. The von Mises stress of dentin, periodontal ligament, alveolar bone, post and core, as well as the periodontal ligament area and threshold limit value were calculated and compared with the findings of traditional crown lengthening models which had been published earlierby our research group. RESULTS: The von Mises stress intensity of modified crown lengthening models were: dentin>post>core>alveolar bone>periodontal ligament. The maximum von Mises stress of dentin(44.37-80.58 MPa)distributed in lingual central shoulder. The periodontal ligament area of the modified crown lengthening surgery was reduced by 6% to 28%, under the same crown-root fracture conditions, the periodontal ligament area of modified crown lengthening models was larger than that of the traditional crown lengthening models. In modified crown lengthening surgery models, the von Mises stress of periodontal ligament of B3L1m, B3L2m, B3L3m models exceeded their limit values, however, the von Mises stress of periodontal ligament of the B2L2c, B2L3c, B3L1c, B3L2c, B3L3c models exceeded their limit values in traditional crown lengthening surgery models. CONCLUSIONS: The modified crown lengthening surgery conserves more periodontal supporting tissues, which facilitates the long-term survival of teeth. The indication of modified crown lengthening surgery is wider than traditional method. The maxillary central incisors with labial fracture at gingival margin level and with palatal fracture at or below the alveolar crest level are not the indication of the crown lengthening surgery.

In silico identification and analysis of phytoene synthase genes in plants.

In this study, we examined phytoene synthetase (PSY), the first key limiting enzyme in the synthesis of carotenoids and catalyzing the formation of geranylgeranyl pyrophosphate in terpenoid biosynthesis. We used known amino acid sequences of the PSY gene in tomato plants to conduct a genome-wide search and identify putative candidates in 34 sequenced plants. A total of 101 homologous genes were identified. Phylogenetic analysis revealed that PSY evolved independently in algae as well as monocotyledonous and dicotyledonous plants. Our results showed that the amino acid structures exhibited 5 motifs (motifs 1 to 5) in algae and those in higher plants were highly conserved. The PSY gene structures showed that the number of intron in algae varied widely, while the number of introns in higher plants was 4 to 5. Identification of PSY genes in plants and the analysis of the gene structure may provide a theoretical basis for studying evolutionary relationships in future analyses.

In silico analysis of gene content in tomato genomic regions mapped to the Ty-2 resistance gene.

Tomato yellow leaf curl virus is one of the main diseases affecting tomato production worldwide. Previous studies have shown that Ty-2 is an important resistance gene located between molecular markers C2_At2g28250 (82.3 cM) and T0302 (89.0 cM), and exhibits strong resistance to tomato yellow leaf curl virus in Asia. In this study, Ty-2 candidate genes were subjected to bioinformatic analysis for the sequenced tomato genome. We identified 69 genes between molecular markers C2_At2g28250 and T0302, 22 of which were disease-related resistant genes, including nucleotide binding site-leucine-rich repeat disease resistance genes, protease genes (protein kinase, kinase receptor, and protein isomerase), cytochromes, and transcription factors. Expressed sequence tag analysis revealed that 77.3% (17/22) of candidate disease-resistance genes were expressed, involving 143 expressed sequence tags. Based on full-length cDNA sequence analysis, 7 candidate genes were found, 4 of which were involved in tomato responses to pathogens. Microarray expression analysis also showed that most candidate genes were involved in the tomato responses to multiple pathogens, including fungi, viruses, and bacteria. RNA-seq expression analysis revealed that all candidate genes participated in tomato growth and development.

Signal transduction pathways in Synechocystis sp. PCC 6803 and biotechnological implications under abiotic stress.

Cyanobacteria have developed various response mechanisms in long evolution to sense and adapt to external or internal changes under abiotic stresses. The signal transduction system of a model cyanobacterium Synechocystis sp. PCC 6803 includes mainly two-component signal transduction systems of eukaryotic-type serine/threonine kinases (STKs), on which most have been investigated at present. These two-component systems play a major role in regulating cell activities in cyanobacteria. More and more co-regulation and crosstalk regulations among signal transduction systems had been discovered due to increasing experimental data, and they are of great importance in corresponding to abiotic stresses. However, mechanisms of their functions remain unknown. Nevertheless, the two signal transduction systems function as an integral network for adaption in different abiotic stresses. This review summarizes available knowledge on the signal transduction network in Synechocystis sp. PCC 6803 and biotechnological implications under various stresses, with focuses on the co-regulation and crosstalk regulations among various stress-responding signal transduction systems.

Enhancement of dissolution of cyclosporine A using solid dispersions with polyoxyethylene (40) stearate.

A solid dispersion containing cyclosporine A (CyA) and polyoxyethylene (40) stearate (PS) was prepared by the solvent-melt method and characterized by powder X-ray diffraction (PXRD), hot-stage microscopy (HSM), scanning electron microscopy (SEM) and dissolution studies. The crystalline peaks of CyA disappeared in the PXRD spectra of solid dispersions but were seen in those of physical mixtures, demonstrating the amorphous state of the drug in solid dispersions. The solubility of CyA in aqueous solutions of PS was increased linearly with increasing amount of PS in water. Dissolution of the drug from solid dispersions and physical mixtures was dramatically enhanced compared to the drug powder alone in water at 37 degrees C.

Expression of IFN-gamma in cerebrovascular endothelial cells from aged mice.

Recently, it has become clear that interferon-gamma (IFN-gamma) plays a role in the central nervous system (CNS) as well as in the immune system. However, the reason for the alteration in IFN-gamma production in the brain with aging remains unknown. In this study, we investigated the expression of IFN-gamma in the brain in terms of both mRNA and protein and compared the expression in young adult brain with that in aged mice. The cerebrum and cerebellum were collected from young adult (8-10 weeks old) and aged (24-26 months old) BALB/c mice, and the expressions of IFN-gamma and IFN-gamma receptor-1 (IFNGR-1) mRNA were examined by RT-PCR. Expression of IFN-gamma mRNA was detected in the brains from aged mice but not in those from young adult mice. However, IFNGR-1 mRNA was expressed in the brains from both young adult and aged mice. Moreover, IFN-gamma levels in the cerebrum and cerebellum from aged mice were detectable by ELISA, but IFN-gamma was undetectable in these tissues from young adult mice. To identify the cellular source of IFN-gamma in the brain of aged mice, immunostaining using antimouse IFN-gamma monoclonal antibody (mAb) was done. Immunoreactivity of IFN-gamma appeared to be located in cerebrovascular endothelial cells, including the choroid plexus of the cerebellum from aged mice. Expression of IFN-gamma and IFNGR-1 was also identified in isolated microvessels from brains. These results suggest that IFN-gamma plays a role in age-associated changes.

Cytokine expression and production by purified Helicobacter pylori urease in human gastric epithelial cells.

Cytokines have been proposed to play an important role in Helicobacter pylori-associated gastroduodenal diseases, but the exact mechanism of the cytokine induction remains unclear. H. pylori urease, a major component of the soluble proteins extracted from bacterial cells, is considered to be one of the virulence factors for the inflammation in the gastric mucosa that is produced in H. pylori infection. However, the response of human gastric epithelial cells to the stimulation of urease has not been investigated. In the present study, we used human gastric epithelial cells in a primary culture system and examined whether H. pylori urease stimulates the gastric epithelial cells to induce proinflammatory cytokines by reverse transcription-PCR and enzyme-linked immunosorbent assay. First, by using peripheral blood mononuclear cells (PBMC) and a gastric cancer cell line (MKN-45 cells), we confirmed the ability of purified H. pylori urease to induce the production of proinflammatory cytokines. Furthermore, we demonstrated that the human gastric epithelial cells produced interleukin-6 (IL-6) and tumor necrosis factor alpha, but not IL-8, following stimulation with purified urease. The patterns of cytokine induction differed among human PBMC, MKN-45 cells, and human gastric epithelial cells. These results suggest that the human gastric epithelial cells contribute to the induction of proinflammatory cytokines by the stimulation of H. pylori urease, indicating that the epithelial cells were involved in the mucosal inflammation that accompanied H. pylori infection.

Intercellular transport of lysosomal acid lipase mediates lipoprotein cholesteryl ester metabolism in a human vascular endothelial cell-fibroblast coculture system.

We present results from studies of human cell culture models to support the premise that the extracellular transport of lysosomal acid lipase has a function in lipoprotein cholesteryl ester metabolism in vascular tissue. Vascular endothelial cells secreted a higher fraction of cellular acid lipase than did smooth muscle cells and fibroblasts. Acid lipase and lysosomal beta-hexosaminidase were secreted at approximately the same rate from the apical and basolateral surface of an endothelial cell monolayer. Stimulation of secretion with NH4Cl did not affect the polarity. We tested for the ability of secreted endothelial lipase to interact with connective tissue cells and influence lipoprotein cholesterol metabolism in a coculture system in which endothelial cells on a micropore filter were suspended above a monolayer of acid lipase-deficient (Wolman disease) fibroblasts. After 5-7 d, acid lipase activity in the fibroblasts reached 10%-20% of the level in normal cells; cholesteryl esters that had accumulated from growth in serum were cleared. Addition of mannose 6-phosphate to the coculture medium blocked acid lipase uptake and cholesterol clearance, indicating that lipase released from endothelial cells was packaged into fibroblast lysosomes by a phosphomannosyl receptor-mediated pathway. Supplementation of the coculture medium with serum was not required for lipase uptake and cholesteryl ester hydrolysis by the fibroblasts, but was necessary for cholesterol clearance. Results from our coculture model suggest that acid lipase may be transported from intact endothelium to cells in the lumen or the wall of a blood vessel. We postulate that delivery of acid hydrolases and lipoproteins to a common endocytic compartment may occur and have an impact on cellular lipoprotein processing.