Tea crude extracts effectively inactivate severe acute respiratory syndrome coronavirus 2.

It is well known that black and green tea extracts, particularly polyphenols, have antimicrobial activity against various pathogenic microbes including viruses. However, there is limited data on the antiviral activity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which emerged rapidly in China in late 2019 and which has been responsible for coronavirus disease 2019 (COVID-19) pandemic globally. In this study, 20 compounds and three extracts were obtained from black and green tea and found that three tea extracts showed significant antiviral activity against SARS-CoV-2, whereby the viral titre decreased about 5 logs TCID50 per ml by 1·375 mg ml-1 black tea extract and two-fold diluted tea bag infusion obtained from black tea when incubated at 25°C for 10 s. However, when concentrations of black and green tea extracts were equally adjusted to 344 µg ml-1 , green tea extracts showed more antiviral activity against SARS-CoV-2. This simple and highly respected beverage may be a cheap and widely acceptable means to reduce SARS-CoV-2 viral burden in the mouth and upper gastrointestinal and respiratory tracts in developed as well as developing countries.

Transcription factor DEC1 is required for maximal experimentally induced periodontal inflammation.

BACKGROUND AND OBJECTIVES: Disruption of transcriptional regulation is a confounding factor associated with a wide range of human inflammatory diseases. To investigate mechanistic links between transcription factor DEC1 and pathways underlying inflammation, wild-type and DEC1 knockout (KO) C57BL/6 mice were treated with Porphyromonas gingivalis (or carboxymethyl cellulose as a control) to induce periodontal inflammation. It provoked an inflammatory response within the oral environment, which showed robust variation in alveolar bone resorption and expression of inflammatory cytokines. MATERIAL AND METHODS: Male DEC1KO mice and their wild-type littermates were used for the experimental periodontitis model. Measurement of alveolar bone resorption, micro-computed tomography, isolation of gingival mononuclear cells (GMCs), flow cytometry and immunohistochemical analysis were used in this study. Human gingival fibroblast cells (HGF-1) were used for DEC1 over-expression and short interference RNA (siRNA) studies and quantitative real-time polymerase chain reaction and western blot analysis were performed. RESULTS: Micro-computed tomography analysis demonstrated that P. gingivalis caused a decrease in bone area of wild-type mice compared with DEC1KO mice. Expression of inflammatory and immune markers in GMCs was significantly decreased in DEC1KO mice after treatment with P. gingivalis. Conversely, interleukin (IL)-4 and IL-10 mRNAs were significantly increased in GMCs isolated from DEC1KO mice. The results show that treatment of DEC1KO mice with P. gingivalis decreased the numbers of CD11b+ F4/80+ and CD4+ RANKL+ T cells. Moreover, expression of CD4, F4/80, RANKL and cathepsin K in inflammatory cell infiltrates was significantly reduced in DEC1KO mice treated with P. gingivalis compared with controls. Furthermore, over-expression of DEC1 in HGF-1 cells increased the expression of IL-1ß and tumor necrosis factor-α mRNAs and their expression levels reached a maximum in response to treatment with lipopolysaccharide. Inhibition of DEC1 by short interference RNA interference suppressed the P. gingivalis-derived lipopolysaccharide-induced expression of IL-1ß, tumor necrosis factor-α and toll-like receptor4. CONCLUSION: These results suggest that transcription factor DEC1 can modulate P. gingivalis-induced periodontitis in the oral mucosa.

Spectrophotometer is useful for assessing vitiligo and chemical leukoderma severity by quantifying color difference with surrounding normally pigmented skin.

BACKGROUND: Acquired skin hypopigmentation has many etiologies, including autoimmune melanocyte destruction, skin aging, inflammation, and chemical exposure. Distinguishing lesions from normally pigmented skin is clinically important to precisely assess disease severity. However, no gold standard assessment method has been reported. We aimed to investigate whether spectrophotometers are useful for assessing vitiligo and rhododendrol (4-(4-hydroxyphenol)-2-butanol) (Rhododenol® )-induced leukoderma disease severity by quantifying skin color. METHODS: Mexameter® MX18 and CM-700d spectrophotometer were used for assessing vitiligo/leukoderma by measuring melanin index, L*a*b* color space, and ΔE*ab value, which represents the color difference between two subjects and is calculated by the values of L*a*b*. RESULTS: MX18 and CM-700d can quantitatively distinguish vitiligo/leukoderma from normally pigmented skin based on melanin index. CM-700d consistently quantified the color of vitiligo/leukoderma lesions and surrounding normally pigmented skin in L*a*b* color spaces and ΔE*ab. ΔE*ab is well correlated with melanin index and clinical appearance. CONCLUSION: ΔE*ab has been frequently used in aesthetic dentistry; however, current study is the first to use it in the measurement of skin color. ΔE*ab seems to be a useful parameter to evaluate the color contrast between vitiligo/leukoderma and surrounding normally pigmented skin and can be used to evaluate disease severity and patient's quality of life.

Surface Properties of Resin Composites and CAD/CAM Blocks After Simulated Toothbrushing.

OBJECTIVES: This study aimed to evaluate the surface gloss, surface roughness, and color change of restorative materials after a three-body wear abrasion. METHODS AND MATERIALS: Four resin composites with different filler particle size (Gracefil Flo [GFF, 0.7 µm], Gracefil LoFlo [GFL, 0.25 µm], Gracefil ZeroFlo [GFZ, 0.15 µm], and Gracefil Putty [GFP, 0.3 µm]), two CAD/CAM resin composite blocks with different filler particle size (Cerasmart 300 [CS3, 0.7 µm] and Cerasmart Prime [CSP, 0.3 µm], GC), and one CAD/CAM lithium disilicate glass-ceramic block (Initial LiSi Block [ILS], GC) as a control were evaluated. Twenty slab-shaped specimens were obtained from each material. Ten specimens were subjected to 80,000 toothbrushing strokes and measured for surface gloss (Gloss Unit, GU), surface roughness (Ra, µm), and color (L*, a*, and b* values) before toothbrushing and at every 20,000 strokes. Color differences (ΔL*, Δa*, Δb*, and ΔE00) before and after toothbrushing were calculated. After 80,000 strokes, abraded surfaces were observed using scanning electron microscopy. The other 10 specimens were measured for Vickers microhardness (VHN). RESULTS: After 80,000 toothbrushing strokes, the mean GU ranged from 60.43 to 16.12 (the highest for ILS and lowest for GFL), and the mean Ra ranged from 0.079 to 4.085 (the lowest for ILS and highest for GFL). At all measuring stages, the calculated ΔE00 values ranged from 0.31 to 0.92 for all materials. The mean VHN ranged from 632.34 to 39.08 (the highest for ILS and lowest for GFZ). The resin composite containing the largest filler particle (GFF) showed significantly lower Ra and higher VHN than other resin composites (GFL, GFZ, and GFP). The CAD/CAM resin composite block containing a smaller filler particle (CSP) retained significantly higher GU than that containing a larger filler particle (CS3). A negative correlation between GU and Ra was detected. CONCLUSIONS: Based on the findings, toothbrush abrasion induced a decrease in GU and an increase in Ra for all resin-based materials tested. Resin-based materials with larger filler size tended to show lower Ra, while resin-based materials with smaller filler size tended to show a smaller reduction in GU. These were more pronounced for light-cure resin composites than for resin composite blocks for CAD/CAM.

Treatment outcomes of proton beam therapy combined with retrograde intra-arterial infusion chemotherapy for locally advanced oral cancer in the elderly.

The aim of this study was to evaluate the therapeutic efficacy and safety of proton beam therapy combined with retrograde intra-arterial infusion chemotherapy in elderly patients with locally advanced oral cancer. Between February 2009 and October 2019, 42 oral cancer patients aged ≥75 years were treated with this therapy. Median age was 80 years (range 75-90 years) and the median follow-up duration was 39 months (range 2-106 months). Of the 42 patients, 34 (81%) were diagnosed with stage IV cancer. The 3-year overall survival, local control, progression-free survival, and disease-specific survival rates were 56%, 69%, 32%, and 67%, respectively. Regarding acute toxicities, grade 3 neutropenia was observed in six patients (14%), anaemia in five (12%), acute kidney injury in one (2%), and oral mucositis in 18 (42%). Late toxicities of grade 3 were observed in seven patients: dysphagia in six (14%) and osteonecrosis of the jaw in one (2%). This study showed that proton beam therapy combined with retrograde intra-arterial infusion chemotherapy was effective for elderly patients with oral cancer, and toxicities were tolerable and manageable. The study findings suggest that this therapy is a potential treatment option for elderly oral cancer patients with difficulty in surgery and systemic chemotherapy.

Flexural Properties and Polished Surface Characteristics of a Structural Colored Resin Composite.

OBJECTIVE: The aim of this study was to determine the flexural properties and surface characteristics of a structural colored resin composite after different finishing and polishing methods, in comparison to those of conventional resin composites. METHODS AND MATERIALS: A structural color resin composite, Omnichroma (OM, Tokuyama Corp, Chiyoda City, Tokyo, Japan), and two comparison resin composites, Filtek Supreme Ultra (FS, 3M, St Paul, MN, USA) and Tetric EvoCeram (TE, Ivoclar Vivadent, Schaan, Liechtenstein), were used. The flexural properties of the resin composites were determined in accordance with the ISO 4049 specifications. For surface properties, 70 polymerized specimens of each resin composite were prepared and divided into seven groups of 10. Surface roughness (Sa), gloss (GU), and surface free energy (SFE) were investigated after the following finishing and polishing methods. Three groups of specimens were finished with a superfine-grit diamond bur (SFD), and three with a tungsten carbide bur (TCB). After finishing, one of the two remaining groups was polished with a one-step silicone point (CMP), and the other with an aluminum oxide flexible disk (SSD). A group ground with SiC 320-grit was set as a baseline. RESULTS: The average flexural strength ranged from 116.6 to 142.3 MPa in the following order with significant differences between each value: FS > TE > OM. The average E ranged from 6.8 to 13.2 GPa in the following order with significant differences between each value: FS > TE > OM. The average R ranged from 0.77 to 1.01 MJ/mm3 in the following order: OM > FS > TE. The Sa values of the OM groups polished with CMP and SSD were found to be significantly lower than those of the other resin composites, regardless of the finishing method. The GU values appeared to be dependent on the material and the finishing method used. The OM specimens polished with SSD showed significantly higher GU values than those polished with CMP. Most of the resin composites polished with SSD demonstrated significantly higher γS values compared to the other groups. Extremely strong negative correlations between Sa and GU in the combined data from the three resin composites and each resin composite and between Sa and γS in the OM specimens were observed; GU showed a strong positive correlation with γS in the same material. CONCLUSION: These findings indicate that both flexural and surface properties are material dependent. Furthermore, the different finishing and polishing methods used in this study were observed to affect the Sa, GU, and SFE of the resin composites.

Prevalence of human papillomavirus in oral gargles and tonsillar washings.

OBJECTIVES: Human papillomavirus (HPV) infection drives carcinogenesis in the oropharynx. No standard sampling or HPV detection methods for evaluating oropharyngeal HPV infection exist. The prevalence of oral HPV infection in Japan is unknown. MATERIALS AND METHODS: We examined 435 healthy Japanese individuals to address whether adding tonsillar washing to oral gargling would improve HPV detection. We compared HPV assessment using GENOSEARCH HPV31 versus nested PCR and direct sequencing. Associations between HPV infection and demographic and behavioral characteristics were examined. RESULTS: Most participants who were HPV-positive based on oral gargles were also HPV-positive based on tonsillar washings: 11 (64.7%) of 17 on nested PCR and 12 (70.6%) of 17 on GENOSEARCH HPV31. Although HPV infection was more prevalent in oral gargles followed by tonsillar washings than in oral gargles alone, the difference was not statistically significant (nested PCR, 4.8% vs. 3.9%, P = 0.46; GENOSEARCH HPV31, 5.3% vs. 3.9%, P = 0.33). The overall agreement between nested PCR and GENOSEARCH HPV31 was 98.6%, with 76.0% positive agreement. The overall prevalence of oral HPV infection in Japan was 5.7% (95% confidence interval, 3.9-8.3%). Men had a significantly higher prevalence of oral HPV infection than women (8.3% vs. 2.6%, P = 0.02). Infection increased with number of lifetime sexual partners (P < 0.001 for trend). CONCLUSION: The oropharynx is probably the major source of HPV-infected cells in oral gargles. Oral gargling could be a standard sampling method for evaluating oropharyngeal HPV infection. GENOSEARCH HPV31 could be an option for oral HPV detection.

Endoscopic sinus surgery for the odontogenic maxillary cysts.

OBJECTIVE: To evaluate the effectiveness and usefulness of transnasal endoscopic surgery for the treatment of odontogenic maxillary cysts. METHODS: Between February 2003 and February 2008, transnasal endoscopic surgery was performed under general anesthesia in 13 patients (male 6 and female 7, 19 to 75 years old) with odontogenic maxillary cysts that extended to the maxillary sinus. Ten patients had a radicular cyst and three patients had a dentigerous cyst. After the resection of anterior edge of the inferior turbinate, the lateral wall of the inferior nasal meatus was opened. Then, the cyst wall of the maxillary sinus was partially or completely removed under the endoscope. RESULTS: The cyst walls were completely removed in five often patients with a radicular cyst and in all three patients with a dentigerous cyst. Five patients with a radicular cyst received partial resection of the cyst wall. The affected teeth could be preserved in seven of ten patients with a radicular cyst and in one of three patients with a dentigerous cyst. There were no complications, and postoperative courses were uneventful. Follow-up period ranged from 11 to 72 months (mean 42 months), and no recurrence has been noted in any of the cases. CONCLUSIONS: Endoscopic transnasal surgery for the odontogenic maxillary cyst is less invasive than conventional dental approach, and most of the affected teeth can be preserved. This technique appears to be a simple and highly effective surgical treatment for the treatment of patients with odontogenic cysts that extend to the maxillary sinus.

Rodent Dental Fluorosis Model: Extraction of Enamel Organ from Rat Incisors.

Chronic fluoride overexposure can cause dental fluorosis. Dental fluorosis is characterized by porous and soft enamel that is vulnerable to erosion and decay. Animal models often contribute to clinical applications by addressing pathogenic questions of disease. To study dental fluorosis, rodent models have been employed because rodent incisors erupt continuously and every stage of enamel development is present along the length of the rodent incisor. Here we present a protocol to induce dental fluorosis in mouse and rat and describe the procedure for extraction of stage specific enamel organ from rat mandibular incisors.

Clinical and Histological Study on Direct Pulp Capping With CO2 Laser Irradiation in Human Teeth.

The study aimed to histologically evaluate wound healing of exposed human pulp on direct pulp capping using super-pulsed CO2 laser preirradiation. In this single-blind clinical trial, 28 third molar teeth of 17 volunteers were randomly capped with either CO2 laser irradiation (n=14) or Dycal (calcium hydroxide cement; n=14) and restored using resin composite. The laser was operated in super-pulsed mode (pulse duration, 0.2 ms; interval, 5.8 ms; 0.003 J/pulse). The irradiation conditions were a power output of 0.5 W, an irradiation time of 15 seconds, repeat mode (10-ms irradiation and 10-ms intervals, for a total beam exposure time of 7.5 seconds), total applied energy of 3.75 J, and an activated air-cooling system. Each tooth was extracted at six or 12 months posttreatment and prepared for histological evaluation. We evaluated the parameters of pulp tissue disorganization, inflammatory cell infiltration, reparative dentin formation (RDF), and bacterial penetration. There were no significant differences between groups for all parameters at each postoperative period (Mann-Whitney U-test, p>0.05). CO2 laser irradiation completely controlled bleeding and exudate from the exposed pulp. The CO2 laser group had a tendency to delay RDF compared with the Dycal group, but 4 of 7 teeth from the CO2 laser group showed a complete dentin bridge at 12 months posttreatment.

Soluble interleukin-1 receptor type II levels in gingival crevicular fluid in aggressive and chronic periodontitis.

BACKGROUND: Interleukin (IL)-1 is closely related to the initiation and progression of periodontal disease. IL-1 levels in the gingival crevicular fluid (GCF) of subjects with periodontitis are higher than those in periodontally healthy controls, and the levels of IL-1 correlate with disease severity. However, soluble IL-1 receptor type II (sIL-1RII), which acts as a decoy receptor for IL-1s, has not been investigated in detail in periodontal disease. The purpose of this study was to measure sIL-1RII levels in the GCF of subjects with chronic or aggressive periodontitis; the correlation between the sIL-1RII levels in GCF and clinical parameters also was examined. METHODS: IL-1beta and sIL-1RII were measured in 64 GCF samples collected from 47 subjects with chronic periodontitis (CP) and 17 subjects with aggressive periodontitis (AgP). The clinical characteristics of each site were recorded at the time of GCF sampling. IL-1beta and sIL-1RII were measured by specific non-cross-reactive enzyme-linked immunosorbent assay. RESULTS: The disease severity was comparable in CP and AgP. IL-1beta was detected in 98% of CP GCF samples and 88% of AgP GCF samples. sIL-1RII was detected in 55% of CP GCF samples and 35% of AgP GCF samples. However, the concentrations of IL-beta and sIL-1RII detected in GCF from subjects with CP or AgP were similar. CONCLUSION: sIL-1RII was detected more often in CP GCF than in AgP GCF, and there was no correlation between GCF sIL-1RII concentration and clinical parameters.

In vitro evaluation of the implant abutment connection sealing capability of different implant systems.

This study sought to evaluate the sealing capability of the implant abutment connection of different dental implant systems. Five Nobel Replace select, Straumann and Intra-lock implants of approximately 4.5 mm diameter with their respective abutments were provided by the manufacturers. A calibration curve was determined by placing toluidine blue (TB) increments of 0.1 microL into 1.5 mL of distilled water and recording its absorbance in a spectrophotometer until reaching 0.7 microL. Then, 0.7 microL of TB was placed in the deepest portion of each implant's internal screw, the abutments were adapted to the implant according to the manufacturer's instructions and the specimens were placed in vials with 1.5 mL of distilled water. Spectrophotometric analysis was performed at 1, 3, 6, 24, 48, 72, 96 and 144 h. Statistical analysis was performed by One-way anova at 95% level of significance. The calibration curve was linear with respect to the TB amount in 1.5 microL distilled water (R(2) = 0.9961). All implant abutment systems presented an increase in absorbance as a function of time. As time elapsed in vitro, significantly higher amounts of TB was released from the Straumann and Nobel Replace Select connection systems (P < 0.0001). Leakage was significant between the groups. Despite controlled torquing, the seal between the implant body and the abutment could not be maintained in all three of the systems tested.

Sinonasal organised haematoma: clinical features and successful application of modified transnasal endoscopic medial maxillectomy.

OBJECTIVE: Although organised haematoma often induces bone thinning and destruction similar to malignant diseases, the aetiology of organised haematoma and the optimal treatment remain unclear. This paper presents the clinical features of individuals with organised haematoma, and describes cases in which a novel modified approach was successfully applied for resection of organised haematoma in the maxillary sinus. METHOD: Pre-operative examination data were evaluated retrospectively. Modified transnasal endoscopic medial maxillectomy was employed. RESULTS: Fourteen patients with organised haematoma were treated. Contrast-enhanced computed tomography showed heterogeneous enhancement in all patients. Eight patients underwent modified transnasal endoscopic medial maxillectomy, without complications such as facial numbness, tooth numbness, facial tingling, lacrimation and eye discharge. Dissection of the apertura piriformis and anterior maxillary wall was not necessary for any of these eight patients. No recurrence was observed. CONCLUSION: Pre-operative examinations can be helpful in determining the likelihood of organised haematoma. Modified transnasal endoscopic medial maxillectomy appears to be a safe and effective method for organised haematoma resection.

DC-STAMP Is an Osteoclast Fusogen Engaged in Periodontal Bone Resorption.

Dendritic cell-specific transmembrane protein (DC-STAMP) plays a key role in the induction of osteoclast (OC) cell fusion, as well as DC-mediated immune regulation. While DC-STAMP gene expression is upregulated in the gingival tissue with periodontitis, its pathophysiological roles in periodontitis remain unclear. To evaluate the effects of DC-STAMP in periodontitis, anti-DC-STAMP-monoclonal antibody (mAb) was tested in a mouse model of ligature-induced periodontitis ( n = 6-7/group) where Pasteurella pneumotropica ( Pp)-reactive immune response activated T cells to produce receptor activator of nuclear factor kappa-B ligand (RANKL), which, in turn, promotes the periodontal bone loss via upregulation of osteoclastogenesis. DC-STAMP was expressed on the cell surface of mature multinuclear OCs, as well as immature mononuclear OCs, in primary cultures of RANKL-stimulated bone marrow cells. Anti-DC-STAMP-mAb suppressed the emergence of large, but not small, multinuclear OCs, suggesting that DC-STAMP is engaged in the late stage of cell fusion. Anti-DC-STAMP-mAb also inhibited pit formation caused by RANKL-stimulated bone marrow cells. Attachment of ligature to a second maxillary molar induced DC-STAMP messenger RNA and protein, along with elevated tartrate-resistant acid phosphatase-positive (TRAP+) OCs and alveolar bone loss. As we expected, systemic administration of anti-DC-STAMP-mAb downregulated the ligature-induced alveolar bone loss. Importantly, local injection of anti-DC-STAMP-mAb also suppressed alveolar bone loss and reduced the total number of multinucleated TRAP+ cells in mice that received ligature attachment. Attachment of ligature induced significantly elevated tumor necrosis factor-α, interleukin-1ß, and RANKL in the gingival tissue compared with the control site without ligature ( P < 0.05), which was unaffected by local injection with either anti-DC-STAMP-mAb or control-mAb. Neither in vivo anti- Pp IgG antibody nor in vitro anti- Pp T-cell response and resultant production of RANKL was affected by anti-DC-STAMP-mAb. This study illustrated the roles of DC-STAMP in promoting local OC cell fusion without affecting adaptive immune responses to oral bacteria. Therefore, it is plausible that a novel therapeutic regimen targeting DC-STAMP could suppress periodontal bone loss.

Appropriate real-time PCR reference genes for fluoride treatment studies performed in vitro or in vivo.

OBJECTIVE: Quantitative real-time PCR (qPCR) is routinely performed for experiments designed to identify the molecular mechanisms involved in the pathogenesis of dental fluorosis. Expression of reference gene(s) is expected to remain unchanged in fluoride-treated cells or in rodents relative to the corresponding untreated controls. The aim of this study was to select optimal reference genes for fluoride experiments performed in vitro and in vivo. DESIGN: Five candidate genes were evaluated: B2m, Eef1a1, Gapdh, Hprt and Tbp. For in vitro experiments, LS8 cells derived from mouse enamel organ were treated with 0, 1, 3 and/or 5mM sodium fluoride (NaF) for 6 or 18 h followed by RNA isolation. For in vivo experiments, six-week old rats were treated with 0 or 100 ppm fluoride as NaF for six weeks at which time RNA was isolated from enamel organs. RNA from cells and enamel organs were reverse-transcribed and stability of gene expression for the candidate reference genes was evaluated by qPCR in treated versus non-treated samples. RESULTS: The most stably expressed genes in vitro according to geNorm were B2m and Tbp, and according to Normfinder were Hprt and Gapdh. The most stable genes in vivo were Eef1a1 and Gapdh. Expression of Ddit3, a gene previously shown to be induced by fluoride, was demonstrated to be accurately calculated only when using an optimal reference gene. CONCLUSIONS: This study identifies suitable reference genes for relative quantification of gene expression by qPCR after fluoride treatment both in cultured cells and in the rodent enamel organ.

The chemical basis for the ferriprotoporphyrin IX-chloroquine complex induced lipid peroxidation.

Ferriprotoporphyrin IX (FPIX) forms a coordination complex with chloroquine, an anti-malarial drug. The FPIX-chloroquine complex strongly promotes the peroxidative cleavage of phospholipid membrane. Iron in the complex is essential for the complex to induce lipid peroxidation. In this paper a more detailed mechanism of the complex promoted lipid peroxidation was investigated. Apotransferrin exhibited no apparent inhibition of the complex evoked lipid peroxidation, indicating no mobilization of iron from the complex. No significant inhibitory effect by superoxide dismutase, catalase and sodium benzoate on the complex induced lipid peroxidative reaction, suggesting little involvement of superoxide anion, hydrogen peroxide and hydroxyl radical in the reaction. Quinine and mefroquine, blood shizontocidal drugs as well as chloroquine, formed a complex with FPIX and each complex more rapidly induced lipid peroxidation than FPIX alone. Primaquine, which is not as effective as quinine or mefroquine on an intraerythrocytic malaria parasite, neither coordinated to FPIX nor promoted lipid peroxidation. The complex formation between FPIX and chloroquine, quinine or mefroquine could play a key role in their anti-malarial actions.

Gene therapy for lung diseases: development in the vector biology and novel concepts for gene therapy applications.

The lung represents an attractive target organ for somatic gene therapy strategy in that, (1) it is easily accessible by vectors, (2) most frequent hereditary disorders, cystic fibrosis (CF) and alpha1-antitrypsin deficiency (alpha1AT), occur in the lung, and (3) carcinoma of the lung is apparently a most common cause of death in humans. To date, approximately 400 clinical protocols for human gene therapy have been approved, and approximately 10% of the protocols target lung diseases such as cystic fibrosis (CF) and lung cancer. Currently available data from some of these human trials have successfully demonstrated that gene transfer to the human lung is possible, and that the strategy of overexpressing exogenous genes for curing or controlling lung diseases is potentially promising. In this manuscript, focusing on gene therapy of lung disorders, we aim to give an overview of the hurdles of current gene transfer strategies to overcome, then and also we aim to review recent, remarkable progresses in the vector biology that are potentially promising to maximize safety and efficiency of gene therapy. In addition, based on the most recent advances in the understanding of the molecular biological aspects of the pathogenesis of lung cancer, asthma, pulmonary fibrosis, and acute lung injury, novel therapeutic strategies of gene therapy for inflammatory and malignant diseases of the lung are discussed.

Purification and characterization of mouse DNA polymerase alpha devoid of primase activity.

A simple method was developed for the isolation of primase-free DNA polymerase-alpha from the DNA polymerase-alpha-primase complex of mouse FM3A cells. The polymerase was separated from primase subunits by chromatography on a single-stranded DNA-cellulose column in the presence of 50% etylene glycol. The primase-free DNA polymerase-alpha contained two polypeptides with molecular masses of 180,000 and 68,000. Analysis of the DNA products with poly(dA)-oligo(dT)10 as template-primer revealed that both primase-free DNA polymerase-alpha and the DNA polymerase-alpha-primase complex predominantly synthesized short DNA with less than 30 nucleotides, but that the DNA polymerase-alpha-primase complex also synthesized some longer DNA with more than 300-400 nucleotides.

Thyrsiferyl 23-acetate is a novel specific inhibitor of protein phosphatase PP2A.

Thyrsiferyl 23-acetate (TF23A), a cytotoxic compound from marine red alga, has been shown to potently and specifically inhibit serine/threonine protein phosphatase 2A (PP2A) with IC50 values of 4-16 microM, depending on the enzyme concentration. TF23A did not affect activity of protein phosphatase 1 (PP1), 2B (PP2B), 2C (PP2C), or protein tyrosine phosphatases (PTP) up to 1 mM. It inhibited PP2A activity in a crude extract of a human T cell line, Jurkat cell, as well as the purified catalytic subunit. Thus, TF23A proved to be a novel useful probe for clearly distinguishing the activity of PP2A from those of the other protein phosphatases in crude cell extracts and identification of cellular processes that are regulated by PP2A.