Natural phytoestrogens such as the isoflavones genistein and daidzein, and the flavones quercetin exhibit anti-cancer properties. This study was purpose to investigate the anti-proliferative effect of phytoestrogens on acute myeloid leukemia (AML) and chronic myeloid leukemia (CML) cells, and their synergistic antileukemic effect in combination with chemotherapeutic drugs. Optimal dosage of genistein, quercetin and in combination with chemicals for leukemia cells were determined by experiments. Cell viability, apoptosis induction and cell cycle arrest were detected by trypan blue staining, MTT assay, optical microscopy, flow cytometry (FCM). The schedule treatment of combination of genistein and chemicals was determined. The results showed that genistein exhibited a dose- and time-dependent inhibitory effect on cell proliferation in NB4 and HL-60 cells, induced apoptosis and cell cycle arrest in G2/M phase. Quercetin had evident inhibitory effect on the proliferation of K562 and K562/A cells. The combination of genistein and chemicals exerted a synergistic effect on cell growth inhibition. In conclusion, this study demonstrated the synergistic antileukemic effect of genistein with chemotherapeutic drugs on leukemic cells. This combination appears to be a new idea for the clinical novel treatment of leukemia.
Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; Cell Proliferation ; drug effects ; Drug Synergism ; Genistein ; pharmacology ; HL-60 Cells ; Humans ; Isoflavones ; pharmacology ; Leukemia, Monocytic, Acute ; pathology ; Leukemia, Myeloid, Acute ; pathology ; Phytoestrogens ; pharmacology ; Quercetin ; pharmacology

Aged ; Female ; Heart Block ; etiology ; Humans ; Ventricular Dysfunction, Left ; etiology

Extensive research in this decade has led to detailed understanding of genetic changes underlying human cancers. Two major tumorigenic events are mutation and amplification of oncogenes and inactivation of tumor suppressor genes. These events then trigger a series of signal transduction cascades, activating expression of downstream genes that control various cellular activities including cell cycle progression, DNA synthesis, programmed cell death, DNA repair, and cell migration. Investigations of these molecular pathways has led to the identification of many targets for therapeutic intervention. Knowledge of the expression patterns and functions of all human genes wil l provide a frame work for future molecular, genetic medicine. During the past ten years, the human genome project has generated an enormous amount of sequencing information, and sequencing of the entire human genome may be completed by the year 2003 (1,2). One can envision that this will irreversibly transform the methodology of medical research and the practice of medicine. The search for new genes, which currently consumes the effort of many talented scientists, will become past history. Additionally, studying one gene at a time will be replaced by studying large number of genes simultaneously(3). Reductionistic approaches to human disease will be replaced by systemic approach. As a prelude to this revolution, tools used for parallel analysis of gene expression in the format of ordered gene arrays have been developed and are under continued expansion. In this technical tip, we will introduce the Atlas Human cDNA Expression Array system developed by Clontech Laboratories, Inc.(4). With this technology, a conventional laboratory can profile the expression of 588 human genes simultaneously in one simple experiment without the using of expensive equipment. We will demonstrate the profiling of 588 genes in a human glioblastoma cell line to exemplify the utility of this technique.
Aptitude ; Cell Cycle ; Cell Death ; Cell Line ; Cell Movement ; DNA ; DNA Repair ; DNA, Complementary* ; Gene Expression ; Genes, Tumor Suppressor ; Genome, Human ; Glioblastoma ; Glioma* ; Human Genome Project ; Humans* ; Oncogenes ; Signal Transduction

Purpose: With advancing endovascular technology and increasing interest in minimally invasive intra-arterial therapies such as stem cell and chemotherapy for cerebral disease, the establishment of a translational model with cerebral circulation accessible to microcatheters is needed. We report our experience catheterizing canine cerebral circulation with microcatheters, present high-resolution angiographic images of the canine vascular anatomy, describe arterial branch flow patterns and provide measurements of canine arterial conduits. Materials and Methods: Angiograms were performed on 10 intact purpose-bred hounds. Angiography, measurements of arterial conduits and catheterization information for intracranial arterial branches were obtained. Results: Selective and superselective cerebral angiography was successful in all subjects. Relevant arterial mean diameters include the femoral (4.64 mm), aorta (9.38 mm), external carotid (3.65 mm), internal carotid arteries (1.6 mm), vertebrobasilar system and Circle of Willis branches. Catheterization of the Circle of Willis was achieved via the posterior circulation in all subjects tested (n=3) and the use of flow directed microcatheters resulted in reduced arterial tree deformation and improved superselection of intracranial vessels. Catheterization of the intracranial circulation was attempted but not achieved via the internal carotid artery (n=7) due to its tortuosity and subsequent catheter related vasospasm. Conclusion The canine cerebral vasculature is posterior circulation dominant. Anterior circulation angiography is achievable via the internal carotid artery, but direct cerebral arterial access is best achieved via the posterior circulation using flow-directed microcatheters. It is feasible to deliver intra-arterial therapies to selective vascular territories within the canine cerebral circulation, thus making it a viable animal model for testing novel intra-arterial cerebral treatments.

Objectives To establish a new transgenic mouse model for determining the function and role of human scavenger receptor A (SR-A) in atherosclerosis in vivo. Methods Human scavenger receptor minigene-driven mouse tie-1 promoter was constructed and confirmed by endonuclease digestion and sequence analysis. Transgenic mice were generated via the microinjection method. PCR and Southern blot were used to screen the positive transgenic mice. RT-PCR and immunohistochemical analysis were used to detect the level and location of human SR-A Ⅰ expression in transgenic mice. The activity of human SR-A Ⅰ was determined by morphologic observation of aortic endothelial cells of transgenic mice under transmission electron microscopy. Results The electrophoresis assay showed the expected 4 fragments of 0.9 kb, 1.1 kb, 1.2 kb and 4.2 kb in the Sma Ⅰ digest and 2 fragments of 0.8 kb and 6.7 kb in Bgl Ⅱ digest of plasmids pTie-1/hSR-A. The fragment sequence of tie-1 promoter and human SR-A cDNA in plasmids pTie-1/hSR-A was correct and no ATG before the translation initiation sites of human SR-A was found by sequence analysis. 561 injected and surviving embryos with the purified human SR-A minigene were implanted into the oviducts of 19 ICR pseudopregnant mice. Among the 54 surviving pups from 13 foster mothers, 7 were identified by PCR and Southern blot analysis. The results of RT-PCR and imrnunohistochemical analysis showed human SR-A was specifically expressed on vascular endothelial cells of the aorta and renal artery, as well as hepatic sinusoidal endothelial cells in transgenic mice. Transmmion electron microscope (TEM) of aorta of transgenic mice showed that a large number of vesicles, multivesicle bodies and swollen mitochondria filled the plasma of endothelial cells. Conclusions A transgenic mouse model with overexpression of human SR-A in endothelial cells was successfully established. The transgene was integrated and transmitted into the chromosome of transgenic mice. Tie-1 promoter controlled the transgene to express in endothelial cells in mice. Pinocytic activity of aortic endothelial cells in transgenic mice was higher than that of C57BL/6J mice. Our studies will provide a new transcgenic model for investigation of atherosclerosis and functions of human SR-A.

Methods: Patients presenting with a spine fracture were diagnosed with AS or DISH at a single tertiary care center between 2010 and 2019. We excluded those who lacked cross-sectional imaging or fractures occurring at spinal segments affected by ankylosis, as well as polytraumatized patients. Patient demographics, injury mechanism, fracture level, neurologic status, treatment, and 1-year mortality were recorded. Computed tomography imaging was reviewed by two independent readers and graded according to the indicated AO Spine Injury Classification System. Differences in fracture severity, treatment method, and mortality were examined using Student t -tests, chi-square tests, and two-proportion Z-tests with significance set to p <0.05. Results: We identified 167 patients with spine fracture diagnosed with AS or DISH. Patients with AS had more severe fractures and more commonly had surgery than patients with DISH (p <0.001). Despite these differences, 1-year mortality did not significantly differ between AS and DISH patients (p =0.14). Conclusions Although patients with AS suffered more severe fractures compared to DISH and more frequently underwent surgery for these injuries, outcomes and 1-year mortality did not differ significantly between the two groups. For patients with ASDs and fractures, outcomes appear similar regardless of treatment modality. Consequently, there may be an opportunity for critical reappraisal of operative indications in ASD and a larger role for nonoperative management in these challenging patients.

Formation of the periodontium begins following onset of tooth-root formation in a coordinated manner after birth. Dental follicle progenitor cells are thought to form the cementum, alveolar bone and Sharpey's fibers of the periodontal ligament (PDL). However, little is known about the regulatory morphogens that control differentiation and function of these progenitor cells, as well as the progenitor cells involved in crown and root formation. We investigated the role of bone morphogenetic protein-2 (Bmp2) in these processes by the conditional removal of the Bmp2 gene using the Sp7-Cre-EGFP mouse model. Sp7-Cre-EGFP first becomes active at E18 in the first molar, with robust Cre activity at postnatal day 0 (P0), followed by Cre activity in the second molar, which occurs after P0. There is robust Cre activity in the periodontium and third molars by 2 weeks of age. When the Bmp2 gene is removed from Sp7(+) (Osterix(+)) cells, major defects are noted in root, cellular cementum and periodontium formation. First, there are major cell autonomous defects in root-odontoblast terminal differentiation. Second, there are major alterations in formation of the PDLs and cellular cementum, correlated with decreased nuclear factor IC (Nfic), periostin and α-SMA(+) cells. Third, there is a failure to produce vascular endothelial growth factor A (VEGF-A) in the periodontium and the pulp leading to decreased formation of the microvascular and associated candidate stem cells in the Bmp2-cKO(Sp7-Cre-EGFP). Fourth, ameloblast function and enamel formation are indirectly altered in the Bmp2-cKO(Sp7-Cre-EGFP). These data demonstrate that the Bmp2 gene has complex roles in postnatal tooth development and periodontium formation.
Actins ; analysis ; Activating Transcription Factor 2 ; genetics ; Age Factors ; Ameloblasts ; pathology ; Amelogenesis ; genetics ; Animals ; Bone Morphogenetic Protein 2 ; genetics ; Cell Adhesion Molecules ; analysis ; Cell Differentiation ; genetics ; Cementogenesis ; genetics ; Dental Cementum ; pathology ; Dental Pulp ; blood supply ; Fluorescent Dyes ; Green Fluorescent Proteins ; Male ; Mice ; Mice, Knockout ; Microvessels ; pathology ; Molar ; growth & development ; Molar, Third ; growth & development ; NFI Transcription Factors ; analysis ; Odontoblasts ; pathology ; Odontogenesis ; genetics ; Periodontal Ligament ; growth & development ; Sp7 Transcription Factor ; Stem Cells ; physiology ; Tooth Root ; growth & development ; Transcription Factors ; genetics ; Vascular Endothelial Growth Factor A ; analysis ; Zinc Fingers ; genetics

OBJECTIVETo investigate the expression and significance of CCL5 in patients with esophageal carcinoma.

METHODSUsing reverse transcriptase polymerase chain reaction (RT-PCR), the expressions of CCL5/CD8/granzyme B/perforin in tumor and corresponding adjacent tissues from esophageal carcinoma patients were examined. Flow cytometry (FACS) was used to detect the percentages of CD8(+) T cells and CCR5(+)CD8(+) T cells in TIL and PBMC from the patients. Transwell assay was performed to study the effect of CCL5 on the migration of T cells in vitro. T test and Spearman correlation analysis were performed.

RESULTSThe mRNA expressions of CCL5 and perforin were 0.348 2 ± 0.300 1 and 0.181 9 ± 0.118 6, respectively, in the tumor samples, while their expressions in adjacent samples were 0.279 6 ± 0.138 0 and 0.118 0 ± 0.109 8, respectively, with no statistically significant differences between them (P > 0.05 for both). The mRNA expressions of CD8 and granzyme B were significantly higher in the tumor tissues than in adjacent tissues (0.464 9 ± 0.300 8 vs. 0.279 0 ± 0.173 4, 0.648 7 ± 0.516 0 vs. 0.469 7 ± 0.259 1; P < 0.05 for both). The relative expression of CCL5 was positively correlated with that of CD8, perforin and granzyme B (r(CD8) = 0.272, P = 0.034; r(perforin) = 0.305, P = 0.026; r(granzymeB) = 0.108, P = 0.012) in the tumor sites. FACS data revealed that the proportions of CD8(+) T cells in TIL and PBMC were (45.86 ± 16.09)% and (34.05 ± 15.07)%, respectively, showing a significant difference (P = 0.022). Similarly, CCR5(+)CD8(+) T cells fraction in TIL (48.12 ± 26.75)% was much higher than that in PBMC (19.53 ± 13.67) % (P < 0.001). Transwell assay showed that CCL5 protein enhanced the migration of T cells, supporting that CCL5 is crucial for CD8(+) T cells recruitment in vivo. Intriguingly, CCL5 expression was down-regulated in advanced patients (stage IIb-IV). The accumulation of CD8(+) T cells and CCR5(+)CD8(+) T cells was strongly reduced in advanced patients, suggesting that CCL5 expression may be involved in the local control of the disease and its reduction may be involved in disease progression.

CONCLUSIONSThe current data indicate the involvement of CCL5 in the regulation of CD8(+) T cell entry into tumor lesions in esophageal carcinoma patients. This process may affect the disease status and potentially as a prognostic factor for cancer patients. Enhancing local CCL5 expression in tumor lesions may represent a novel strategy in esophageal cancer therapy.

CD8-Positive T-Lymphocytes ; Chemokine CCL5 ; metabolism ; Disease Progression ; Esophageal Neoplasms ; metabolism ; Flow Cytometry ; Humans ; Leukocytes, Mononuclear ; Lymphocytes, Tumor-Infiltrating

BACKGROUND/AIMS: Nausea, an unpleasant symptom of diabetic gastroparesis (DMGP), has been reported to be alleviated by needleless transcutaneous electrical acupuncture (TEA). Our study was designed to utilize electroencephalography (EEG) and electrogastrography (EGG) recordings to investigate the central and peripheral responses of TEA in the treatment of nausea in DMGP patients. METHODS: Eleven DMGP subjects underwent simultaneous EEG and EGG testing while grading the severity of nausea following 30-minute intervals of: (1) baseline, (2) visual stimulation (VS) to provoke more nausea, (3) active VS together with TEA, and (4) TEA alone, and a final 15-minute recording without any intervention. RESULTS: The nausea score was increased to 5.9 ± 1.5 with VS (P < 0.05, vs 3.5 ± 1.0 at baseline), then reduced to 3.5 ± 1.2 with VS plus TEA, and to 2.5 ± 1.3 with TEA alone, while it continued at a score of 2.9 ± 1.0 post TEA (all significant, P < 0.05, vs VS without TEA). The mean percentage of normal gastric slow waves was decreased to 60.0 ± 5.7% with VS (P < 0.05, vs 66.6 ± 4.5% at baseline), then improved to 69.2 ± 4.8% with VS plus TEA, and maintained at 70 ± 3.6% with TEA alone. During initial VS, EEG signals showed right inferior frontal activity as the prominent finding, but during VS with TEA, left inferior frontal activity predominated. CONCLUSIONS: In DMGP, TEA improves gastric dysrhythmia and ameliorates nausea. TEA treatment of nausea provoked by VS resulted in a change of dominance from right to left inferior frontal lobe activity. These data provide new understandings of peripheral and central mechanisms for nausea, and potential future directions for DMGP treatment approaches.
Acupuncture* ; Electroencephalography ; Frontal Lobe ; Gastroparesis* ; Humans ; Nausea* ; Ovum ; Photic Stimulation ; Tea