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Bisphenol A (BPA) is an endocrine-disrupting chemical, widely used to produce polycarbonate plastic. Carnosic acid (CA) is a rosemary diterpene with an anti-obesity effect. In this study, we investigated the anti-adipogenic effect of CA in BPA-treated 3T3-L1 preadipocytes and C57BL/6 J mice. In vitro experiments showed that CA inhibited lipid accumulation by BPA in 3T3-L1 preadipocytes. CA displayed anti-adipogenic effects through the downregulation of differentiation and adipogenesis-related proteins, along with the upregulation of lipolytic protein and SIRT1/FoxO1 pathway. In vivo experiments, mice treated with BPA exhibited an increase in body weight gain and epididymal adipose tissue mass when compared to the control group. CA treatment improved the epididymal adipose tissue mass induced by BPA. CA and rosemary extract (RE) treatment ameliorated dyslipidemia in BPA-treated mice. We further showed that CA and RE exerted anti-adipogenesis effects in liver tissues of BPA-treated mice via increasing SIRT1, FoxO1, and ATGL proteins and decreasing FAS and aP2 proteins. Moreover, SIRT1 inhibitor sirtinol blocked CA to increase SIRT1, FoxO1, FAS, and aP2 proteins, decrease Ac-FoxO1 protein, and reduce lipid accumulation in BPA-treated cells. These findings indicated that CA and RE could reverse BPA-induced lipid accumulation by regulating adipocyte differentiation, adipogenesis, and lipolysis through SIRT1/FoxO1 pathway., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Kaoliang is a refreshing fragranced type of Chinese spirits with slight apple fragrance that comes from ethyl acetate (EA). Special aromas are produced by esterification microorganisms, which affect the taste and quality of the wine. In this study, new yeast strains were isolated from yellow water, a by-product during fermentation process. Meanwhile, the optimal culture condition was determined for its growth and EA production. Three new strains, Kazachstaniaexigua, Candida humilis and Saccharomyces cerevisiae were identified from yellow water. Among these strains, S. cerevisiae S5 was the new and dominant strain. Results from response surface methodology showed that S. cerevisiae S5 produced 161.88 ppm of EA, in the medium with 4.91% yeast extract, 9.82% peptone, and 20.91% glucose after 96 hours of cultivation at 27.53°C. GC analysis showed that aroma compounds, such as EA, isoamyl acetate and 2-phenylethanol increased from the sample of optimal condition when compared to the one from initial fermentation condition., Competing Interests: The authors have declared that no competing interests exist.

Background: Pseudoachalasia is a rare diagnosis manifested by clinical and physiologic symptoms of achalasia, with alternative etiology for outflow obstruction. While malignancy is a frequent cause of pseudoachalasia, prior surgical intervention especially surgery involving the esophagogastric junction, may result in a misdiagnosis of achalasia., Case Presentation: We present a case of a 70 year-old male with dysphagia and weight loss after undergoing a Billroth I and Nissen fundoplication several decades ago. His preoperative studies suggested achalasia and he was therefore referred for an endoscopic myotomy. However, careful interpretation of all the data and intra-operative findings revealed a classic mechanical and functional obstruction requiring takedown of his prior wrap., Conclusions: Individualized interpretation of preoperative studies in the setting of prior foregut surgery is critical to appropriate diagnosis and intervention. This case highlights the significance of endoscopic findings and features of high-resolution manometry specific to pseudoachalasia, which contrasts with classical features of achalasia.

This in vitro study compared the ultrastructural quality of the apical seal achieved with Resilon/Epiphany and gutta-percha/AH Plus. Single-rooted extracted human teeth were prepared using a crown-down technique, debrided with NaOCl and EDTA, and obturated with either Resilon/Epiphany or gutta-percha/AH Plus. They were examined for gaps along canal walls using SEM, and for apical leakage using a transmission electron microscopy (TEM). SEM revealed both gap-free regions, and gap-containing regions in canals filled with both materials. TEM revealed the presence of silver deposits along the sealer-hybrid layer interface in Resilon/Epiphany, and between the sealer and gutta-percha in the controls. It is concluded that a complete hermetic apical seal cannot be achieved with either root filling materials.

Aim: To analyze the immunological properties and biological activity of a monoclonal antibody (mAb) against CD3 molecule(yCD3), and to observe the tumor-suppressive activity of CD3AK cells in vitro and in vivo., Methods: FCM was used to test the specificity of yCD3 and the immunological phenotype and cytokine production of CD3AK. 3H-TdR assay was used to measure the transformation of lymphocytes activated by yCD3. LDH assay was used to analyze the cytotoxic activity of CD3AK in vitro. Mice bearing tumors were used to observe the anti-tumor effect of CD3AK cells., Results: yCD3 could bind specifically to T cells. 5 microg yCD3 could competitively inhibit 70% of standard anti-CD3 antibody to bind with CD3 molecules on cell membrane. 8 microg/L of yCD3 stimulated the proliferation of peripheral blood lymphocytes, which could be further boosted by IL-2 or anti-CD28 antibodies. Among activated CD3AK cells, CD3+, CD8+, and CD25+ cells increased. IL-2 and IFN-gamma producing CD3+ cells were also increased, to 3.29- and 2.47- fold, respectively, under the co-stimulation of anti-CD28 antibody. When the ratio of effective cells and target cells was 80:1, 57.54% target cells were killed. As compared with control, the percent of tumor inhibition in CD3AK cells treated tumor-bearing mice was 33.17%, and the inhibition rate of lung metastasis was 39.70%. The CD3AK cells treatment was more effective when combined with LAK cells., Conclusion: yCD3 could activate T cells and significantly induce the tumor-suppressive activity of CD3AK cells in vitro and in vivo, which lays a foundation for adoptive immunotherapy against tumors in clinical medicine.

Aim: To construct and express anti-human CD3 chimeric antibody., Methods: The genes of variable regions of the light chain (V(L)) and heavy chain (V(H)) were cloned respectively into the expression vectors (V(L) Express, V(H) Express), and co-transfected into COS-7 cells. Expression level of the chimeric antibody in culture supernatant was detected by ELISA. The antibody was purified through protein A affinity chromatography and identified by Western blot. Binding activity of the chimeric antibody to the antigen was determined by FACS. Biological activity of the chimeric antibody was determined by mixed T-lymphocyte culture test., Results: The expression vectors were constructed and the anti-human CD3 chimeric antibody was expressed and purified successfully. Western blot showed that the purified antibody was human-mouse chimeric antibody. FACS result showed that the chimeric antibody had antigen-binding activity. Mixed T-lymphocyte culture test showed that the chimeric antibody could suppress proliferation of T lymphocytes., Conclusion: The anti-human CD3 chimeric antibody has been constructed and expressed successfully, which lays the foundation for its further study.

Objectives: This study tested the hypothesis that single-bottle total-etch adhesives are effective in reducing dentine permeability under in vivo conditions., Methods: Crown preparations on vital human teeth were performed under local analgesia as part of the treatment plan for prosthetic rehabilitation. Four single-bottle adhesives (Single Bond, 3M ESPE; Excite DSC, Ivoclar Vivadent; Prime and Bond NT Dual-Cure, Dentsply DeTrey and One-Step, Bisco Inc.) were applied to the cut dentine after acid-etching. Polyvinyl siloxane impressions were taken, using an ultra-low viscosity impression material, of the smear layer-covered dentine before applying the adhesives to deep vital dentine, and after adhesive placement. Additional impressions were taken of the adhesive-sealed dentine following the removal of the provisional prostheses after a 7-10 day period. Epoxy resin replicas of the crown preparations were examined with scanning electron microscopy to evaluate the extent of dentinal fluid transudation during pre-bonded, immediately bonded and post-bonded periods., Results: Dentinal fluid transudation from localised areas that were close to the dental pulp was universally observed from all epoxy resin replicas, irrespective of the adhesive employed. The transudation of dentinal fluid from the control smear layer-covered dentine of each crown preparation was comparatively mild when compared to the extent that was observed after total-etching and application of the single-bottle adhesive. Dentinal fluid droplets were specifically located over the surface of the adhesive layer. Continuous transudation of dentinal fluid occurred even after the removal of the provisional prostheses, and was considerably more profuse in subjects who opted for the fitting of the permanent prostheses without the use of a local anaesthetic solution., Conclusion: Single-bottle adhesives, because of their lack of a comparatively more hydrophobic bonding resin layer, behave as permeable membranes after polymerisation. They permit the continuous transudation of dentinal fluid and do not provide a hermetic seal in vital deep dentine. Although the relatively slow rate of diffusion of dentinal fluid is unlikely to result in post-operative cold sensitivity, it may interfere with the optimal polymerisation of dual-cured or auto-cured composites or resin cements in both direct and indirect restorations.

Aim: To develop a Fab antibody against platelet GPIIb/IIIa., Methods: A mouse anti-human GPIIb/IIIa monoclonal antibody (mAb) P140 was selected by the indirect immunofluorescence assay (IFA) and platelet aggregation inhibition test. The Fd and light chain genes were cloned from the hybridoma cells secreting the mAb P140. The genes of P140 Fab were inserted into plasmid p3MH to construct recombinant expression plasmid p3MH/P140kappa-Fd. After digestion with the restriction enzyme, the recombinant plasmid was transformed into E.coli XLI-Blue. The expressed product was purified by TALON metal affinity resin. Purified P140 Fab was characterized by SDS-PAGE, ELISA, Western blot and platelet aggregation inhibition test., Results: SDS-PAGE analysis showed that relative molecular mass (M(r)) of P140 Fab was 47x10(3). The results of ELISA, Western blot and platelet aggragation inhibition test indicated that P140 Fab could specifically bind to platelet and inhibit platelet aggragation in dose-dependent manner. The mean value of IC(50) was 16.85 mg/L., Conclusion: A soluble anti-platelet GPIIb/IIIa antibody P140 Fab was prepared successfully, which lays the foundation for further clinical application.

One-step self-etch adhesives behave as permeable membranes after polymerization, permitting water to move through the cured adhesives. We hypothesize that osmotic blistering occurs in bonded enamel when these adhesives are used without composite coupling. Tooth surfaces from extracted human premolars were bonded with 5 one-step self-etch adhesives. They were immersed in distilled water or 4.8 M CaCl(2), and examined by stereomicroscopy, field-emission/environmental SEM, and TEM. Water blisters were observed in bonded enamel but not in bonded dentin when specimens were immersed in water. They collapsed when water was subsequently replaced with CaCl(2). Blisters were absent from enamel in specimens that were immersed in CaCl(2) only. Water trees were identified from adhesive-enamel interfaces. Osmotic blistering in enamel is probably caused by the low water permeability of enamel. This creates an osmotic gradient between the bonded enamel and the external environment, causing water sorption into the interface.

In this study, to investigate the effect on expression of IL-2 in lymphocytes from bone marrow and peripheral blood of normal donors after they were mobilized by G-CSF in allo-BMT, 7 normal donors bone marrow and peripheral blood were harvested before and after G-CSF administration. The separated lymphocytes were measured by FCM after they were stained intracellularly by anti-IL-2, and their expressions of IL-2 were compared. The degree of aGVHD in patients after bone marrow transplantation was evaluated clinically, and it was compared with the status of aGVHD of 15 patients whose donors didn't receive G-CSF administration in our department, and 2 groups of patients are comparable in age, types of diseases and status of donors. The results showed that the expression of IL-2 in lymphocytes in 7 G-CSF mobilized donors decreased significantly after G-CSF administration and more severe aGVHD than grade II didn't develop in these recipient patients, and comparing with 15 patients received the bone marrow from donors who didn't receive G-CSF, the incidence of aGVHD decreased. It is suggested that the expression of IL-2 in lymphocytes was influenced by donors' G-CSF administration, and it is likely that thereby reduces the incidence of aGVHD in patients after BMT.

CD20 is a specific antigen expressed on normal and neoplastic B cells exclusively. Recent researches showed that in B cell leukemia, CD20 was over-expressed. Therefore monoclonal antibody (McAb) to CD20 may be of clinical value in diagnosis and treatment of some leukemias and lymphomas. In this study, the full length gene of CD20 cDNA were cloned from total RNA of Raji cells, inserted into an eukaryotic expression vector pcDNA3.1, forming a recombinant plasmid pcDNA3.1/CD20. NIH-3T3 cells were transfected with pcDNA3.1/CD20 and selected with G418 for the transfected cells. Alkaline phosphatase against alkaline phosphatase assay(APAAP) experiments showed that the selected cells could express the human CD20 onto its surface. Balb/c mice were immunized with CD20( ) NIH-3T3 cells once three weeks for 3 shuts. Indirect immunofluorescence experiments were done with the Raji cells and the serum of the immunized mice, and the results showed that the spleen of the immunized mice could be used to prepare the McAb to CD20.

Using cell surface markers and a panel of monoclonal antibodies, 515 cases of acute lymphoblastic leukemia (ALL) were immunophenotyped. T cell type ALL (T-ALL), non-T cell type ALL (Non-T-ALL) including common ALL (C-ALL), Null-ALL and B cell type ALL (B-ALL) were found. These major subtypes of ALL were further divided according to their phenotypes in detail. It was noticed that the phenotypes of these subtypes of ALL reflected basically the phenotypes of normal T or B cells at various differentiation stages or certain population of lymphocytes. The diagnosis of cell lineage was more precise when based on immunophenotyping than morphological description. The combination of morphological and immunological classification can improve the diagnosis of acute leukemias. In addition, it was observed that the immunophenotyping was relevant to clinicopathologic features, responses to therapy and prognosis of ALL patients. The incidences of major subtypes of ALL, the age distribution of ALL subsets and male sex bias with T-ALL in Chinese are discussed.

The immunophenotype of leukemicblasts from 111 patients with T-ALL or T-NHL were further examined by using a panel of standardized McAbs of CD nomenclature to human leukocyte differentiation antigens. Four major subsets of T-ALL were defined: pre T-ALL, immature T-ALL (I), common T-ALL (II) and mature T-ALL (III), with the percentages 20.7%, 20.7%, 20.7% and 37.0% respectively. In addition there was a case with M-T acute hybrid leukemia. Some of the clinical features of the patients with T-ALL and T-NHL were compared. It was found that male predominance, older age, higher leukocyte count, lower platelet level, relative higher hemoglobin level and increased incidence of extramedullary involvement, including hepatomegaly, splenomegaly and lymphadenopathy were alike for all subsets of T-ALL cases. However, the average white cell level and incidence of lymphadenopathy in the pre T-ALL subset significantly differed from those in other subsets. The correlation of immunophenotype with morphologic characterization was also discussed in this paper.

Primary hepatocellular carcinoma(s) (PHC) induced in rats by N-2-fluorenylacetamide (AAF) or diethylnitrosamine (DEN) were examined for their bioelectric potential (EP). N-2-Fluorenylacetamide-induced PHC differed from age-matched normal livers by a mean of -12.9, which is significant at p less than 0.001. They also differed from non-PHC-bearing surrounding liver by -4.75 mV, significant at p less than 0.05. Diethylnitrosamine-induced PHC differed from normal liver by -13.5 mV and from background liver by -9.95 mV, both significant at p less than 0.001. To determine the contribution of cell division to these differences, regenerating livers were examined. The levels of mixed function oxidases in PHC and in regenerating livers reported in the literature appeared to correlate with the measured EP. We, therefore, studied the effects of chemical inhibitors and an inducer of this enzyme system. The alterations of mixed function oxidase were demonstrated to parallel those of EP.

High potassium concentrations altered the morphology and the ability to grow in soft agar in 6m2 cells, a clone of rat kidney cells infected with a temperature-sensitive mutant of Moloney sarcoma virus. Approximately 60% of cells exhibited normal morphology in the presence of 94.8 mM potassium in isotonic medium at the temperature permissive for transformation, whereas 100% were normal at 72 mM potassium in hypertonic media. A significant reduction of growth in soft agar was also induced with these conditions. However, the synthesis ratio of virus-specified transforming protein to marker viral protein was not altered. Na+K+-ATPase might play a role in this reverse-transformation process.

Normal rat kidney cells (NRK) infected with the temperature-sensitive (ts) transformation mutant of Moloney murine sarcoma virus yielded a clone of cells, 6m2, that exhibited a transformed morphology at 33 degrees C and a normal morphology at 39 degrees C. Transmembrane potential (Em) was measured fluorometrically using a cyanine dye diS-C3-(5). Fluorescence was inversely correlated with Em. Cells at 33 degrees C had lower Em. Em changes were recorded within 15 minutes of temperature shift from 33 degrees C to 39 degrees C in both directions, increasing in the 33 degrees C to 39 degrees C direction and decreasing in the 39 degrees C to 33 degrees C direction. Uninfected NRK cells when shifted under the same condition exhibited small fluorescence changes in the 33 degrees C to 39 degrees C direction. Shifting from 39 degrees C to 33 degrees C resulted in Em changes similar to those in 6m2 cells. Also studied was a cell line infected with a spontaneous revertant of the ts mutant, designated 54-5A4; it was transformed at both temperatures. Shifting from 33 degrees C to 39 degrees C in both directions yielded small changes. Transmembrane potential changes in 6m2 cells precede other transformation-specific changes that occur after a temperature shift.

Free flow electrophoresis measurements have been used to determine the surface charge density of normal rat kidney (NRK) cells and a clone of NRK, designated as 6m2, that exhibit a transformed phenotype at 33 degrees C and a non-transformed phenotype at 39 degrees C. A clone of 6m2, designated 54-5A4, which is transformed at both 33 degrees C and 39 degrees C was also studied. A surface charge density of -1.42 microC/cm2 was obtained for the NRK and non-transformed 6m2 cells at 39 degrees C, whereas at 33 degrees C values of -1.85 and -1.78 microC/cm2 were determined for the transformed 6m2 and 54-5A4 cells, respectively. It was found that 72% of the increased charge that appeared on the transformed 6m2 cells compared with the non-transformed 6m2 cells was RNAase sensitive. The time-dependent decrease in surface charge that accompanied the shift of the 6m2 cells from their transformed to non-transformed state was found to mirror the increase in transmembrane potential previously reported using a fluorescent dye technique, and was also comparable to the reported temporal changes in their morphology and virally-coded protein content.

K+ at high concentrations (52-72 mM hypertonic KCl) has been reported to induce reverse transformation in the 6m2 cell, which is a clone of normal rat kidney cells (NRK) infected with a temperature-sensitive transformation virus. When exposed to high K+, 6m2 cells grown at the permissive temperature (33 degrees C) exhibit normal morphology and reduced soft agar growth, characteristics of cells grown at nonpermissive temperature (39 degrees C). In the current study, flattening of cells and rearrangement of surface microvilli were demonstrated by scanning electron microscopy to occur within 6 hr of exposure to high K+, similar to the effect of temperature shift to 39 degrees C. Exposure to K+ resulted in a 90% inhibition of P85gag-mos-associated serine kinase activity within 5 min, with a subsequent reduction of up to 75% of the synthesis of this protein. These alterations in the putative transforming protein were similar to those induced by temperature shift and were considered to be the basis for retrotransformation. The cell microtubular system and F-actin cables were affected more slowly by K+ than by a temperature shift to 39 degrees C. The former did not achieve the fine reticulum network seen in NRK cells until 72 hr later, but the latter remained aberrant. The effect on the enzyme might be mediated by alteration in phosphorylation, but the mechanism by which kinase inactivation induces retrotransformation is not yet known.