Biomedical subjects

Zhao Wang

Publications and source records attributed to Zhao Wang.

At least 37 records · Page 2Linked to original sources

PubMed2005

Patterns of neoplastic foci and lymph node micrometastasis within the mesorectum.

BACKGROUND AND AIMS: Local recurrence after rectal cancer surgery is conceived to result from microscopically incomplete resection. We aimed to investigate the patterns of mesorectal neoplastic foci, and examined the involvement and micrometastasis of lymph nodes. METHODS: Observation of large tissue slice and analysis of tissue microarray were integrated in the pathological study of 31 total mesorectal excision (TME) specimens. RESULTS: Altogether, 349 mesorectal neoplastic foci were examined from 18 specimens. Almost 33% of them were in the outer layer of mesorectum. Concerning position of primary tumor, ipsilateral neoplastic foci were significantly more than contralateral neoplastic foci. Distal mesorectal spread was found in four patients with the distance ranging from 1 to 3.5 cm. Four specimens were diagnosed to have circumferential margin involved. Nine hundred seventy-two lymph nodes were harvested with 128 involved by tumor. No significant difference in occurrence of micrometastasis was observed among tumors of different stage. CONCLUSIONS: Combination of large tissue slice and tissue microarray provided a more detailed method in studying the metastasis of rectal cancer. Complete excision of the mesorectum with fascia propria circumferentially intact is essential. Circumferential margin involvement and micrometastasis suggested that tumor spread may go beyond the scope of a single TME procedure.

Adenocarcinoma↗

PubMed2005

[Comparison of therapeutic effects of low-dose versus high-dose interferon alpha-2b treatment on chronic myelocytic leukemia: a prospective randomized study].

OBJECTIVE: To compare the therapeutic effects of low-dose and high-dose interferon alpha-2b (IFN) treatment on chronic myelocytic leukemia (CML). METHODS: A real-time quantitative reverse transcriptase PCR (RQ-PCR) method was established to detect the fusion gene bcr-abl expression, thereby studying the reduction of leukemic cells. Thirty newly diagnosed CML patients, 21 males and 9 females, aged 14 - 69, were treated with hydroxyurea to keep the white blood cell count less than 20 x 10(9)/L, and then randomized into 2 groups: high-dose IFN group receiving IFN alpha-2b 5MIU 6 times per week for 3 - 6 months and low-dose IFN group receiving IFN alpha-2b 3MIU every other day for 3 - 6 months. Bone marrow was collected every month to Real-time PCR was used to detect the expression of bcr-abl mRNA. Mononuclear cells were isolated and RNA was extracted to detect the expression of fusion gene bcr-abl and a control gene GAPDH. The results were reported as the number of bcr-abl copies/GAPDH copy. RESULTS: The established real-time quantitative PCR method could detect the bcr-abl molecules as low as 50 copies. The intra-assay coefficient of variation (CV) was less than 5% and the inter-assay CV was 5.13%. The median bcr-abl fusion gene expression level of 30 CML patients before IFN therapy was 0.098 (range: 0.010 - 5.799). The bcr-abl expression level decreased by 19.37% and 24.86% in the low-dose and high-dose IFN groups respectively after 3 months' therapy. No significant difference was observed between the two groups (P = 0.398). Relatively more side effects were observed in the high-dose IFN group than in low-dose group. CONCLUSION: RQ-PCR is a reliable method to monitor CML therapy by analyzing fusion gene bcr-abl expression. There is a difference in bcr-abl fusion gene expression levels among the newly diagnosed patients, and low-dose IFN is as effective as high-dose IFN in reducing bcr-abl expression but with less side effects.

Adolescent↗

PubMed2005

Cloning and expression analysis of gonadogenesis-associated gene SPATA4 from rainbow trout (Oncorhynchus mykiss).

Gonadogenesis is a complicated process which involves multi-gene interactions. A rainbow trout (Oncorhynchus mykiss) gene spermatogenesis associated 4 (SPATA4) was cloned and characterized from adult rainbow trout testis. The cDNA sequence of rainbow trout SPATA4 contains an open reading frame of 1, 081 nucleatides encoding a putative protein of 259 amino acids. The putative protein from rainbow trout shares a 76.8% homology with zebrafish SPATA4. No trans-membrane regions or signal peptide were detected using bioinformatics methods. Subcellular localization analysis revealed that rainbow trout SPATA4 was a nuclear protein with highest possibility (39.1%). Multi-tissue reverse transcriptase PCR (RT-PCR) was performed to examine the distribution of rainbow trout SPATA4 in eleven organs of adult rainbow trout. The result demonstrated that this gene express specifically in testis and slight amount of expression was detected in ovary. Further analysis of SPATA4 characterization and function in rainbow trout may provide insight into the understanding of gonadogenesis process.

Amino Acid Sequence↗

PubMed2005

Differences in platelet endothelial cell adhesion molecule-1 expression between peripheral circulation and pancreatic microcirculation in cerulein-induced acute edematous pancreatitis.

AIM: To investigate the changes of platelet endothelial cell adhesion molecule-1 (PECAM-1) expression on polymorphonuclear leukocytes (PMNs) in peripheral circulation and pancreatic microcirculation in cerulein-induced acute edematous pancreatitis (AEP). METHODS: Fifty Wistar rats were randomly divided into control group (n = 10) and AEP group (n = 40). A model of AEP was established by subcutaneous injection of cerulein 5.5 and 7.5 mug/kg at 0 and 1 h after the beginning of experiment respectively. PECAM-1 expression on PMNs from splenic vein and inferior vena cava was determined by RT-PCR at mRNA level and determined by flow cytometry at protein level. RESULTS: In experimental rats, an increased PECAM-1 mRNA expression was seen from 4 to 8 h of AEP in peripheral circulation (0.77+/-0.25%, 0.76+/-0.28%, 0.89+/-0.30%, 1.00+/-0.21%), while in pancreatic microcirculation, expression decreased from 2 h and reached the lowest level at 6 h of AEP (0.78+/-0.29%, 0.75+/-0.26%, 0.62+/-0.28%, 0.66+/-0.20%). There were significant differences at 8-h time point of AEP between peripheral circulation and pancreatic microcirculation (1.00+/-0.21% vs 0.66+/-0.20%, P<0.05). Meanwhile, the difference at protein level was also found. CONCLUSION: A reverse expression of PECAM-1 on PMNs was found between peripheral circulation and pancreatic microcirculation, suggesting that inhibition of PECAM-1 expression may improve the pathological change of AEP.

Acute Disease↗

PubMed2005

Pituitary adenylate cyclase activating-peptide and its receptor antagonists in development of acute pancreatitis in rats.

AIM: Pituitary adenylate cyclase activating-peptide (PACAP) is a late member of the secretin/glucagon/vasoactive intestinal peptide (VIP) family of brain-gut peptides. It is unknown whether PACAP takes part in the development of acute pancreatitis and whether PACAP or its antagonists can be used to suppress the progression of acute pancreatitis. We investigated the actions of PACAP and its receptor antagonists in acute pancreatitis on rats. METHODS: Acute pancreatitis was induced in rats with caerulein or 3.5% sodium taurocholate. The rats were continuously infused with 5-30 microg/kg PACAP via jugular vein within the first 90 min, while 10-100 microg/kg PACAP6-27 and (4-Cl-D-Phe6, Leu17) VIP (PACAP receptor antagonists) were intravenously infused for 1 h. Biochemical and histopathological assessments were made at 4 h after infusion. Pancreatic and duodenal PACAP concentrations were determined by enzyme-linked immunosorbent assay (ELISA). Chinese ink-perfused pancreas was fixed, sectioned and cleared for counting the functional capillary density. RESULTS: PACAP augmented caerulein-induced pancreatitis and failed to ameliorate sodium taurocholate-induced pancreatitis. ELISA revealed that relative concentrations of PACAP in pancreas and duodenum were significantly increased in both sodium taurocholate- and caerulein-induced pancreatitis compared with those in normal controls. Unexpectedly, PACAP6-27 and (4-Cl-D-Phe6, Leu17) VIP could induce mild acute pancreatitis and aggravate caerulein-induced pancreatitis with characteristic manifestations of acute hemorrhagic/necrotizing pancreatitis. Functional capillary density of pancreas was interpreted in the context of pancreatic edema, and calibrated functional capillary density (calibrated FCD), which combined measurement of functional capillary density with dry weight/wet weight ratio, was introduced. Hyperemia or congestion, rather than ischemia, characterized pancreatic microcirculatory changes in acute pancreatitis. CONCLUSION: PACAP may take part in the pathogenesis of acute pancreatitis in rats. The two PACAP receptor antagonsits might act as partial agonists. Calibrated functional capillary density can reflect pancreatic microcirculatory changes in acute pancreatitis.

Acute Disease↗

PubMed2005

Pathological study of distal mesorectal cancer spread to determine a proper distal resection margin.

AIM: Local recurrence after curative surgical resection for rectal cancer remains a major problem. Several studies have shown that incomplete removal of cancer deposits in the distal mesorectum contributes a great share to this dismal result. Clinicopathologic examination of distal mesorectum in lower rectal cancer was performed in the present study to assess the incidence and extent of distal mesorectal spread and to determine an optimal distal resection margin in sphincter-saving procedure. METHODS: We prospectively examined sepecimens from 45 patients with lower rectal cancer who underwent curative surgery. Large-mount sections were performed to microscopically observe the distal mesorectal spread and to measure the extent of distal spread. Tissue shrinkage ratio was also considered. Patients with involvement in the distal mesorectum were compared with those without involvement with regard to clinicopathologic features. RESULTS: Mesorectal cancer spread was observed in 21 patients (46.7%), 8 of them (17.8%) had distal mesorectal spread. Overall, distal intramural and/or mesorectal spreads were observed in 10 patients (22.2%) and the maximum extent of distal spread in situ was 12 mm and 36 mm respectively. Eight patients with distal mesorectal spread showed a significantly higher rate of lymph node metastasis compared with the other 37 patients without distal mesorectal spread (P = 0.043). CONCLUSION: Distal mesorectal spread invariably occurs in advanced rectal cancer and has a significant relationship with lymph node metastasis. Distal resection margin of 1.5 cm for the rectal wall and 4 cm for the distal mesorectum is proper to those patients who are arranged to receive operation with a curative sphincter-saving procedure for lower rectal cancer.

Adult↗

PubMed2005

Methyl protodioscin induces G2/M arrest and apoptosis in K562 cells with the hyperpolarization of mitochondria.

Methyl protodioscin is a furostanol bisglycoside with antitumor properties. The present study investigated its effects on human chronic myelogenous leukemia K562 cells. Cell cycle analysis showed that methyl protodioscin caused distinct G2/M arrest, with the appearance of polyploidy population. The levels of cyclin B1 decreased, whereas Cdc2 kept at a steady level. Subsequent apoptosis after G2/M blockage was demonstrated through DNA fragmentation and the annexin V staining assay. Methyl protodioscin induced a biphasic alteration (i.e. an early hyperpolarization, followed by depolarization) in mitochondrial membrane potential of K562 cells. The transient decline of intracellular Ca2+ concentration was observed at early stage. The generation of reactive oxygen species was also detected. The anti-apoptotic Bcl-x(L) transiently increased and then decreased. And the pro-apoptotic Bax was markedly up-regulated. Taken together, these data demonstrated that methyl protodioscin inhibits K562 cell proliferation via G2/M arrest and apoptosis, with mitochondrial hyperpolarization and the disruption of Ca2+ homeostasis playing important roles.

Apoptosis↗

PubMed2005

Cloning and characterization of zebra fish SPATA4 gene and analysis of its gonad specific expression.

The spermatogenesis associated 4 gene (SPATA4, previously named TSARG2) was first cloned in human tissues and was reported to be a candidate spermatocyte apoptosis-related gene that is expressed specifically in testis. Analysis of SPATA4 expression and regulation in zebra fish may provide insight into the understanding of the complicated process of gonadogenesis. In this study, we cloned and characterized the SPATA4 gene from zebra fish (Danio rerio), which is homologous to human and mouse SPATA4. Zebra fish SPATA4 consists of six exons separated by five introns, as all SPATA4 genes in vertebrates. A promoter region was predicted using homologous blast and cloned for further study, and possible transcription factors were analyzed in this region. The putative protein encoded by this gene was analyzed using bioinformatics methods. Multi-tissue RT-PCR results demonstrated that the zebra fish SPATA4 gene is expressed specifically in testis and slightly in ovary. Analysis of the SPATA4 sequence and its spatial expression pattern indicate that this gene is highly conserved and may play an important role in the process of zebra fish gonadogenesis.

Amino Acid Sequence↗

PubMed2005

Noninvasive intraocular pressure measurements in mice by pneumotonometry.

PURPOSE: To develop a reliable, noninvasive, continuous, and easily implemented system for measuring intraocular pressure (IOP) in mice. METHODS: Pneumotonometry was adapted for measurement of mouse IOP. Measurements were compared with those obtained with the servo-null micropipette system (SNMS) and with direct anterior chamber cannulation. Heart rate was monitored by the precordial pulse, EKG, or tail pulse in anesthetized mice. The characteristic ocular hypotensive response to mannitol was assessed as an additional validation of the METHOD RESULTS: Measurements of IOP obtained using pneumotonometry agreed closely with values measured by SNMS or by direct cannulation. IOP oscillations were synchronous with the heart rate, with a coherence peak between them of approximately 2 Hz, equal to the pulse frequency. Hypertonic mannitol reduced IOP from 13.7 +/- 0.9 mm Hg by 7.7 +/- 0.7 mm Hg after 15 minutes. CONCLUSIONS: Pneumotonometry is a reliable and noninvasive method for the measurement of IOP in mice and may permit comparisons of IOP to hemodynamic factors. This system is simpler and more adaptable for glaucoma research than previously reported methodologies for measuring IOP in mice.

Animals↗

PubMed2005

Tolvaptan administration does not affect steady state amiodarone concentrations in patients with cardiac arrhythmias.

BACKGROUND: Tolvaptan, a nonpeptide selective vasopressin receptor (V2) antagonist, is in development for the treatment of congestive heart failure and hyponatremia. Tolvaptan is primarily metabolized via CYP3A4. This study was conducted to determine the extent of the pharmacokinetic interaction between tolvaptan and steady state amiodarone, an antiarrhythmic drug commonly prescribed for patients with congestive heart failure and a known inhibitor of other drugs metabolized by CYP3A4. METHODS: This was a multicenter, open-label, 1-arm, 3-period, sequential treatment study conducted in 11 men (10) and women aged 49 to 80 years. They were primarily Caucasian (20) subjects, with a history of cardiac arrhythmias who were otherwise healthy. Subjects were to have been on oral amiodarone maintenance therapy of 200 mg/day for at least 10 months. All subjects took 200 mg amiodarone once daily on each study day; on days 3 and 4, they were also coadministered 30 and 90 mg of tolvaptan, respectively. The plasma concentrations of amiodarone and its metabolite desethylamiodarone were determined for 24 hours postdose on days 2, 3, and 4, tolvaptan concentrations were determined for 24 hours postdose on days 3 and 4. RESULTS: As determined by the ratio of the geometric means and 90% confidence intervals (0.5 to 2.0) for the maximal plasma concentration and the area under the curve during the dosing interval for both amiodarone and desethylamiodarone, tolvaptan coadministration had no effect on either amiodarone and desethylamiodarone disposition, as all the geometric mean ratios (amiodarone + tolvaptan [30 or 90 mg] vs amiodarone alone) were approximately 1. CONCLUSION: Tolvaptan coadministration does not alter steady-state amiodarone or desethylamiodarone concentrations. Tolvaptan concentrations did not appear to be different from historical controls. The most frequently reported adverse event was polyuria (15 of 21 subjects for amiodarone + 30 mg tolvaptan); an expected outcome due to the known potent aquaretic action of tolvaptan. The combination of amiodarone and tolvaptan was well tolerated.

Aged↗

PubMed2005

Overexpression of QM induces cell differentiation and mineralization in MC3T3-E1.

It has been reported that QM was highly expressed by cells isolated from epiphyseal cartilage as opposed to proliferative chondrocytes. In vitro investigation of the expression of QM revealed higher QM expression in nonmineralizing osteoblast and pericyte cultures as compared with mineralizing cultures. These evidences suggest that QM may play an essential role in cell differentiation before mineralization. However, our research results showed that QM overexpression in MC3T3-E1 enhanced cell differentiation and mineralization. In this study, alkaline phosphatase (ALP) activity and nodule mineralization were increased in MC3T3-E1 from QM overexpression cultures relative to normal expression QM cultures. RT-PCR revealed upregulation of the marker genes type I collagen, ALP, osteocalcin, osterix and BMP-2 and a slight decrease of a negative regulator osteopontin. These results suggest that the increasing of QM expression could stimulate osteoblast differentiation and mineralization in MC3T3-E1.

3T3 Cells↗

PubMed2005

[Effects of analogs of pituitary adenylate cyclase activating polypeptide on acute pancreatitis in rats].

OBJECTIVE: To examine the effects of analogs of pituitary adenylate cyclase activating polypeptide (PACAP)--PACAP6-27 (10, 100 microg/kg) and (4-Cl-D-Phe6, Leu17)VIP (10, 100 microg/kg) on the pancreata of normal rats and on the development of experimental acute pancreatitis. METHODS: Male Wistar rats were allocated into normal control groups, experimental acute pancreatitis groups and PACAP analog intervention groups. Acute pancreatitis was induced with s.c. cerulein and intraductal sodium taurocholate; PACAP analogs were infused intravenously immediately after pancreatitis induction. Pancreatic morphology was observed at 4 h, and serum amylase, pancreatic water content and PACAP contents were measured. RESULTS: It was found that PACAP6-27 induced pancreatic edema, inflammatory cell infiltration, and elevation of serum amylase [(1464.33 +/- 265.6)-(1692.17 +/- 312.18)] IU/L vs (520.8 +/- 163.27) IU/L of control, P < 0.05); that PACAP6-27 aggravated vacuolization of pancreatic acinar cells in cerulein-induced pancreatitis with hemorrhage and fatty and parenchymal necrosis; and that the pathological changes of cerulein plus 100 microg/kg PACAP group were similar to those of sodium taurocholate-induced pancreatitis. Pancreatic hemorrhage, vacuolization of acinar cells and parenchymal necrosis in sodium taurocholate-induced pancreatitis were worsened by PACAP6-27. (4-Cl-D-Phe6, Leu17)VIP had similar effects. ELISA showed that pancreatic and duodenal levels of PACAP were increased in cerulein- and sodium taurocholate-induced pancreatitis. CONCLUSION: PACAP6-27 and (4-Cl-D-Phe6, Leu17)VIP could induce mild pancreatitis and aggravate experimental acute pancreatitis. PACAP probably plays a role in the pathogenesis of acute pancreatitis.

Acute Disease↗

PubMed2005

[Monitoring all-trans-retinoic acid-induced differentiation of HL-60 cells by Fourier-transform infrared spectroscopy].

This study reports a new in vitro analytical method, based on Fourier-transform infrared (FTIR) spectroscopy, tomonitor themyeloid differentiation process in human myeloblast leukemia HL-60 cells induced by all-trans-retinoic acid (ATRA). The alteration of characteristic bands was identified in the differentiated cells, arising from proteins, lipids, carbohydrates and nucleic acids. Besides the changes in lipid content and plasma membrane fluidity, the most striking changes were observed in the region of nucleic acids and carbohydrates. The authors speculate that the glycosylation and phosphorylation of proteins and the hydrogen-bonding of nucleic acids were involved in differentiation. The spectral parameters were correlated with the differentiation index, as determined by NBT reduction assay. These results suggest that FTIR spectroscopy can be used to monitor the differentiation process of HL-60 cells.

Antineoplastic Agents↗

PubMed2005

[Study with large slice technique and tissue microarray on lymph node involvement and micro-metastasis of rectal cancer].

OBJECTIVE: To study lymph node involvement and micro-metastasis of rectal cancer with large slice technique and tissue microarray. METHODS: Large slice technique, combined with tissue microarray,was used in pathologic study of 31 patients after total mesorectal excision (TME) for rectal cancer. RESULTS: Nine hundred and ninety- two lymph nodes were harvested and 148 were positive. More than 40% of positive lymph nodes were located in the outer layer of the mesorectum and in the same side of the mesorectum as the primary tumor was. Circumferential margin involvement was observed in 12 cases and correlated with the numbers of metastatic lymph nodes (Beta =1.166, P=0.041). Micrometastasis was found in 9 cases with negative pathological lymph nodes, but not correlated with tumor differentiation and stage (P> 0.05). CONCLUSION: Large slice technique combined with tissue microarray facilitates the detection of lymph node involvement and micrometastasis. There is a predominance of lymph node metastasis in the outer layer and the same side of the mesorectum. Micrometastasis can be discovered in different stages of rectal cancer.

Adult↗

PubMed2005

[Systems biology and traditional Chinese medicine (TCM) modernizatin].

Systems biology is a new field in biology that aims at system-level understanding of biological systems. Based on the remarkable progress from molecular biology, systems biology focuses its research on investigating the biological systems to form their entirety. Considering the trend of TCM modernization and the coherence on the point of holism between systems biology and TCM, a comprehensive way for the research of TCM modernization was referred, which involves a new concept, componentomics.

Drug Combinations↗

PubMed2004

Microscopic spread of low rectal cancer in regions of the mesorectum: detailed pathological assessment with whole-mount sections.

BACKGROUND AND AIMS: The aim was to investigate the regional spread of microscopic tumor nodules in the mesorectum of patients with low rectal cancer, and to provide further pathological evidence for optimal procedure selection of radical resection for rectal cancer. PATIENTS AND METHODS: Sixty-two patients with low rectal cancer underwent low anterior resection and total mesorectal excision (TME). Surgical specimens were sliced transversely on serial embedded blocks at 2.5-mm intervals, and stained with hematoxylin and eosin. On whole-mount sections the mesorectum was divided into three regions: the outer region of the mesorectum (ORM), the middle region of the mesorectum (MRM), and the inner region of the mesorectum (IRM). Microscopic metastatic foci were investigated for metastatic mesorectal region, frequency, types, involvement of the lymphatic system, and correlation with the primary tumor. RESULTS: Microscopic spread of the tumor was observed in 38.7% of patients (24 out of 62) and that in the ORM was observed in 25.8% of the patients (16 out of 62). Microscopic tumor foci spread in the circumferential resection margin (CRM) occurred in 6.5% of the patients (4 out of 62), and distal mesorectum (DMR) involvement was detected in 6.5% (4 out of 62), with the spread extending to within 3 cm from the lower margin of the tumor. CONCLUSIONS: The results of the present study support the theory that complete excision of the mesorectum without destruction of the ORM is essential for surgical management of low rectal cancer, and an optimal DMR clearance resection margin should be no less than 4 cm. Four patients with microscopic tumor nodule spread in the ORM observed in the study suggested that microscopic metastases exist in pelvic lateral areas and in the mesorectum simultaneously, indicating the significance of preoperative and/or postoperative radiochemotherapy.

Adenocarcinoma↗

PubMed2004

Microscopic spread of low rectal cancer in regions of mesorectum: pathologic assessment with whole-mount sections.

AIM: To assess the microscopic spread of low rectal cancer in mesorectum regions to provide pathological evidence for the necessity of total mesorectal excision (TME). METHODS: A total of 62 patients with low rectal cancer underwent low anterior resection and TME, surgical specimens were sliced transversely on the serial embedded blocks at 2.5 mm interval, and stained with hematoxylin and eosin (HE). The mesorectum on whole-mount sections was divided into three regions: outer region of mesorectum (ORM), middle region of mesorectum (MRM) and inner region of mesorectum (IRM). Microscopic metastatic foci were investigated microscopically on the sections for the metastatic mesorectal regions, frequency, types, involvement of lymphatic vessels and correlation with the original rectal cancer. RESULTS: Microscopic spread of the tumor in mesorectum and ORM was observed in 38.7% (24/62) and 25.8% (16/62) of the patients, respectively. Circumferential resection margin (CRM) with involvement of microscopic metastatic foci occurred in 6.5% (4/62) of the patients, and distal mesorectum (DMR) involved was 6.5% (4/62) with the spread extent within 3 cm of low board of the main lesions. Most (20/24) of the patients with microscopic metastasis in mesorectum were in Dukes C stage. CONCLUSION: Results of the present study support that complete excision of the mesorectum without destruction of the ORM is essential for surgical management of low rectal cancer, an optimal DMR clearance resection margin should be no less than 4 cm, further pathologic assessment of the regions in extramesorectum in the pelvis is needed.

Adenocarcinoma↗

PubMed2004

Diosgenin induces cell cycle arrest and apoptosis in human leukemia K562 cells with the disruption of Ca2+ homeostasis.

PURPOSE: Diosgenin is a steroidal sapogenin with estrogenic and antitumor properties. In order to elucidate the mechanism of its antiproliferative activity, we investigated its effects on the cell cycle and apoptosis in human chronic myelogenous leukemia K562 cells. METHODS: Cell viability was assessed via an MTT assay. Apoptosis was investigated in terms of nuclear morphology, DNA fragmentation, and phosphatidylserine externalization. Cell cycle analysis was performed via PI staining and flow cytometry (FCM). Western blotting and immunofluorescence methods were used to determine the levels of p53, cell cycle-related proteins and Bcl-2 family members. FCM was also used to estimate the changes in mitochondrial membrane potential (MMP), intracellular Ca2+ concentration and reactive oxygen species (ROS) generation. RESULTS: Cell cycle analysis showed that diosgenin caused G2/M arrest independently of p53. The levels of cyclin B1 and p21Cip1/Waf1 were decreased, whereas cdc2 levels were increased. Subsequent apoptosis was demonstrated with the dramatic activation of caspase-3. A dramatic decline in intracellular Ca2+ concentration was observed as an initiating event in the process of cell cycle arrest and apoptosis, which was followed by the hyperpolarization and depolarization of MMP. Generation of ROS was observed in the progression of apoptosis. The antiapoptotic Bcl-2 and Bcl-xL proteins were downregulated, whereas the proapoptotic Bax was upregulated. CONCLUSIONS: Diosgenin inhibits K562 cell proliferation via cell cycle G2/M arrest and apoptosis, with disruption of Ca2+ homeostasis and mitochondrial dysfunction playing vital roles.

Apoptosis↗