A Phase 3, Randomized, Controlled Trial Evaluating the Efficacy and Safety of Ropeginterferon Alfa-2b in Patients with Moderate COVID-19.

INTRODUCTION: Ropeginterferon alfa-2b is a novel mono-pegylated proline-interferon. This clinical study aimed to evaluate its antiviral efficacy of ropeginterferon alfa-2b against SARS-CoV-2 infection. METHODS: This is a multicenter, randomized, open-label study. Adult patients with confirmed SARS-CoV-2 infection with initial cycle threshold (Ct) value < 30 and symptom onset within 4 days were enrolled. Eligible patients were randomized in a 2:1 ratio to receive a single 250-µg dose of ropeginterferon alfa-2b subcutaneously plus standard of care (SOC) or to receive SOC alone. The primary endpoint was the proportion of patients with a negative RT-PCR result for SARS-CoV-2 or discharged from the hospital before Day 8. Change in clinical status based on the World Health Organization (WHO) clinical progression scale and pulmonary infiltrations through chest radiograph were also evaluated. RESULTS: A total of 132 patients were enrolled and treated with study medication. Higher percentages of patients who achieved Ct ≥ 30 or were discharged from the hospital were observed on Day 8 and every other time point of assessment, i.e., Days 5, 11, 15, and 22, in the ropeginterferon alfa-2b group compared to the SOC alone group. However, the difference was statistically significant on Day 11 but not on Day 8. The primary endpoint was not met. The ropeginterferon alfa-2b group showed a higher improvement rate in lung infiltration on Day 5 (27.6% vs. 0.0%, p = 0.0087) and a higher improvement rate in WHO clinical progression scores on Day 8 (69.4% vs. 35.3%, p = 0.03) than those in the SOC group. No ropeginterferon alfa-2b-related serious adverse event was observed. CONCLUSION: Our data show that ropeginterferon alfa-2b with SOC shortened the duration of SARS-CoV-2 shedding compared with SOC alone. In addition, ropeginterferon alfa-2b as an additional therapy could be beneficial by improving lung infiltration.

Sciatica caused by spinal epidural abscess as the initial clinical presentation of colon cancer: a rare case report and review of literature.

BACKGROUND: Colorectal cancer is one of the most frequently diagnosed forms of cancer, and it is associated with several common symptoms and signs such as rectal bleeding, altered bowel habits, abdominal pain, anemia, and unintentional weight loss. Sciatica, a debilitating condition in which the patient experiences paresthesia and pain in the dermatome of associated lumbosacral nerve roots or sciatic nerve distribution, is not considered one of these. Here we present a case of colorectal cancer manifesting symptoms of sciatica alone. CASE PRESENTATION: A 68-year-old male presented with progressive lower back pain radiating to his left thigh and calf over L5/S1 dermatome. Sciatica was suspected and initially underwent conservative treatment with analgesics. However, the symptoms progressed and MRI revealed an epidural abscess surprisingly. Surgical debridement was performed and pus culture isolated Streptococcus gallolyticus. Based on the strong association of S. gallolyticus with colorectal cancer, the presence of this pathogen prompted further tumor evaluation, even in the absence of the typical symptoms and signs. This investigation ultimately leads to the diagnosis of sigmoid adenocarcinoma. CONCLUSIONS: Although rare, sciatica caused by S. gallolyticus infection of the spinal epidural space may serve as the initial presentation of colorectal cancer. Physicians should be aware of the strong association between S. gallolyticus and colorectal cancer. Based on what we currently know about the condition; a thorough systematic assessment of occult neoplasia for patients with S. gallolyticus infection is recommended.

Comparable clinical outcomes with same-day versus rapid initiation of antiretroviral therapy in Taiwan.

OBJECTIVES: WHO has recommended same-day antiretroviral therapy (SDART) initiation since 2017; however, higher attrition rates were noted in developing countries. METHODS: We included newly diagnosed people with HIV (PWH) from 2018 to 2022 at 18 hospitals around Taiwan. SDART initiation was defined as starting ART on the same day of HIV diagnosis and rapid initiation as starting ART within 14 days of diagnosis. A composite unfavorable outcome was defined as death after 30 days of diagnosis, loss to follow-up (LTFU), or virologic failure or rebound at 12 months. RESULTS: At 12 months, PWH on SDART initiation and those on rapid ART initiation showed similar rates of engagement in care with plasma HIV-1 RNA <50 copies/mL (87.5% vs 87.7%) and composite unfavorable outcome (7.7% vs 7.7%). PWH aged >30 years were less likely to have LTFU (aHR 0.44, 95% CI 0.28-0.70). PWH aged >30 years (aHR 0.59, 95% CI 0.41-0.85) and gay, bisexual, and men who have sex with men (GBMSM) (aHR 0.50, 95% CI 0.32-0.79) were less likely to have composite unfavorable outcomes. CONCLUSIONS: SDART and rapid ART initiation resulted in comparable clinical outcomes and viral suppression rates. PWH aged >30 years and GBMSM were less likely to have unfavorable outcomes.

Treatment responses to integrase strand-transfer inhibitor-containing antiretroviral regimens in combination with short-course rifapentine-based regimens for latent tuberculosis infection among people with HIV.

BACKGROUND: Real-world experience with combinations of short-course rifapentine-based regimens and integrase strand-transfer inhibitor (InSTI)-containing antiretroviral therapy (ART) in management of latent tuberculous infection (LTBI) is limited among people with HIV (PWH). METHODS: From August 2019 to October 2022, PWH receiving 3 months of weekly rifapentine plus isoniazid (3HP) or 1 month of daily rifapentine plus isoniazid (1HP) in combination with ART were included. The primary outcome was virologic response within 12 months after LTBI treatment, and the secondary outcomes included treatment completion rate and safety of LTBI regimens. RESULTS: During the study period, 479 PWH (94.6% male; median age, 43 years) were included: 142 received 1HP and bictegravir (BIC)-containing regimens (1HP/BIC group), 46 1HP and dolutegravir (DTG)-containing regimens (1HP/DTG group), 38 3HP and BIC-containing regimens (3HP/BIC group), 214 3HP and DTG-containing regimens (3HP/DTG group), 17 1HP and other ART regimens (1HP/others group), and 22 3HP/other ART regimens (3HP/others group). In the intention-to-treat analysis, the proportions of PWH maintaining plasma HIV-1 RNA <200 copies/mL within 12 months after LTBI treatment completion were 96.5% (1HP/BIC), 100% (1HP/DTG), 100% (3HP/BIC), 95.8% (3HP/DTG), 100% (1HP/others), and 100% (3HP/others). The overall completion rates were >80% for all treatment groups, whereas >50% of the included PWH experienced any adverse event. LTBI regimens and ART combinations were not associated with virologic response and completion rate. CONCLUSION: Combinations of short-course rifapentine-based regimens and InSTI-containing ART maintained viral suppression for most PWH within 12 months of LTBI treatment completion with low rates of grade 3 or higher adverse events.

Assessment of the detection accuracy of SARS-CoV-2 rapid antigen test in children and adolescents: An updated meta-analysis.

BACKGROUND: Real-time and appropriate antigen tests play a pivotal role in preventing severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection. However, a previous meta-analysis reported that the antigen test had lower sensitivity for the detection of SARS-CoV-2 in children. To provide a comprehensive evaluation of diagnostic efficiency, we performed an updated meta-analysis to assess the detection accuracy of SARS-CoV-2 antigen tests stratified by days after symptom onset and specimen type in children and adolescents. METHODS: We comprehensively searched for appropriate studies in the PubMed, Embase, and Cochrane Library databases. Studies on the diagnostic accuracy of antigen tests for SARS-CoV-2 in children and adolescents were included. The relevant data of the included studies were extracted to construct a 2 × 2 table on a per-patient basis. The overall sensitivity and specificity of the SARS-CoV-2 antigen tests were estimated using a bivariate random-effects model. RESULTS: Seventeen studies enrolling 10 912 patients were included in the present meta-analysis. For the detection accuracy of SARS-CoV-2 antigen tests, the meta-analysis generated a pooled sensitivity of 77.9% (95% confidence interval [CI]: 67.3%-85.8%) and a pooled specificity of 99.6% (95% CI: 98.9%-99.8%). The subgroup analysis of studies that examined antigen tests in symptomatic participants ≦7 days after symptom onset generated a pooled sensitivity of 79.4% (95% CI: 47.6%-94.2%) and a pooled specificity of 99.4% (95% CI: 98.2%-99.8%). Another subgroup analysis of studies that evaluated nasal swab specimens demonstrated a pooled sensitivity of 80.1% (95% CI: 65.0%-89.7%) and a pooled specificity of 98.5% (95% CI: 97.3%-9.2%). CONCLUSION: Our findings demonstrated that the antigen test performed using nasal swab specimens exhibited high sensitivity for the detection of SARS-CoV-2 within 7 days after symptom onset. Therefore, antigen testing using nasal swabs may be effective in blocking SARS-CoV-2 transmission in children.

Polyhydramnios as a sole ultrasonographic finding for detecting fetal hemolytic anemia caused by anti-c alloimmunization.

OBJECTIVE: The prenatal course of a rare case with fetal anemia caused by maternal anti-c alloimmunization was reported. CASE REPORT: A 39-year-old female with anti-c and anti-E antibodies against red cells had previously experienced a stillbirth. At her present pregnancy, titers of maternal antibodies and fetal middle cerebral artery peak systolic velocity (MCA-PSV) were frequently monitored to investigate the severity of fetal hemolytic anemia. Rather than manifesting as an increase in MCA-PSV, the anemic fetus was delivered at 32 weeks and one day of gestation with a sole presentation: polyhydramnios. Neonatal hospitalization course were compatible with hemolytic anemia. The baby was discharged at 48 days of age. CONCLUSION: This case illustrated the complexities of dealing with maternal red cell alloimmunization during pregnancy and the limitations of noninvasive diagnostic modalities for detecting fetal anemia, and highlighted that obstetricians should refer all available clinical parameters in order to offer appropriate perinatal care.

Bisdemethoxycurcumin suppresses human brain glioblastoma multiforme GBM 8401 cell migration and invasion via affecting NF-κB and MMP-2 and MMP-9 signaling pathway in vitro.

Human glioblastoma (GBM) is one of the common cancer death in adults worldwide, and its metastasis will lead to difficult treatment. Finding compounds for future to develop treatment is urgent. Bisdemethoxycurcumin (BDMC), a natural product, was isolated from the rhizome of turmeric (Curcuma longa), which has been shown to against many human cancer cells. In the present study, we evaluated the antimetastasis activity of BDMC in human GBM cells. Cell proliferation, cell viability, cellular uptake, wound healing, migration and invasion, and western blotting were analyzed. Results indicated that BDMC at 1.5-3 µM significantly decreased the cell proliferation by MTT assay. BDMC showed the highest uptake by cells at 3 h. After treatment of BDMC at 12-48 h significantly inhibited cell motility in GBM 8401 cells by wound healing assay. BDMC suppressed cell migration and invasion at 24 and 48 h treatment by transwell chamber assay. BDMC significantly decreased the levels of proteins associated with PI3K/Akt, Ras/MEK/ERK pathways and resulted in the decrease in the expressions of NF-κB, MMP-2, MMP-9, and N-cadherin, leading to the inhibition of cell migration and invasion. These findings suggest that BDMC may be a potential candidate for the antimetastasis of human GBM cells in the future.

Hepatitis B virus X gene impacts on the innate immunity and immune-tolerant phase in chronic hepatitis B virus infection.

BACKGROUND AND AIMS: The immunologic features involved in the immune-tolerant phase of chronic hepatitis B (CHB) virus (HBV) infection are unclear. The hepatitis B virus X (HBx) protein disrupts IFN-ß induction by downregulating MAVS and may destroy subsequent HBV-specific adaptive immunity. We aimed to analyse the impacts of genetic variability of HBx in CHB patients on the immune-tolerant phase during long-term follow-up. METHODS: Children with CHB in the immune-tolerant phase were recruited and followed longitudinally. HBx gene sequencing of infecting HBV strains was performed, and the effects of HBx mutations on the immune-tolerant phase were assessed. Restoration of the host immune response to end the immune-tolerant phase was investigated by immunoblotting, immunostaining, ELISA and reporter assays of MAVS/IFN-ß signalling in liver cell lines, patient liver tissues and the HBV plasmid replication system. RESULTS: A total of 173 children (median age, 6.92 years) were recruited. Patients carrying HBx R87G, I127V and R87G + I127V double mutations exhibited higher cumulative incidences of immune-tolerant phase breakthrough (p = .011, p = .006 and p = .017 respectively). Cells transfected with HBx R87G and I127V mutants and pHBV1.3-B6.3 replicons containing the HBx R87G and I127V mutations exhibited statistically increased levels of IFN-ß, especially under poly(I:C) stimulation or Flag-MAVS cotransfection. HA-HBx wild-type interacted with Flag-MAVS and enhanced its ubiquitination, but this ability was diminished in the R87G and I127V mutants. CONCLUSIONS: HBx suppresses IFN-ß induction. R87G and I127V mutation restored IFN-ß production by preventing MAVS degradation, contributing to curtailing the HBV immune-tolerant phase in CHB patients.

Interleukin-6 Test Strip Combined With a Spectrum-Based Optical Reader for Early Recognition of COVID-19 Patients With Risk of Respiratory Failure.

The COVID-19 pandemic has had a globally devastating impact. This highly contagious virus has significantly overburdened and undermined medical systems. While most infected patients experience only mild symptoms, those who are severely affect require urgent medical interventions and some develop acute respiratory failure and require mechanical ventilation. The broad and potentially deadly impact of infection underscores the critical need for early recognition, especially for those at risk for respiratory failure. Those who are severely impacted and at high risk for respiratory failure have been found to present high levels of serum cytokines, such as interleukin-6 (IL-6). Timely diagnosis and management of those at risk for respiratory failure is crucial. Measurement of IL-6 may provide a means for distinguishing such patients. Currently, most serum IL-6 detection relies on the use of laboratory-based conventional enzyme-linked immunosorbent assays. Although some rapid assays have been developed recently, they need to be conducted by specific technicians in central laboratory settings with advanced and expensive equipment. In this study, we propose an IL-6 test strip combined with a spectrum-based optical reader for early recognition of COVID-19-infected patients at imminent risk of acute respiratory failure requiring mechanical ventilator support. For our analyses, clinical demographic data and sera samples were obtained from three medical centers, and test strip specificity and detection performance were analyzed. This would help healthcare personnel stratify the risk of respiratory failure and provide prompt, and suitable management.

Demethoxycurcumin Promotes Macrophage Cell Population and Phagocytosis in WEHI-3 Cell-generated Leukemia BALB/c Mice In Vivo.

BACKGROUND/AIM: Demethoxycurcumin (DMC), one of the components of curcuminoids, has antitumor activities in many human cancer cells and is known to induce apoptosis in human leukemia cells. However, there are no reports showing the effects of DMC on the immune response in leukemia mice in vivo. Herein, we evaluated the impact of DMC on immune responses in WEHI-3-generated leukemia mice in vivo. MATERIALS AND METHODS: Fifty male BALB/c mice were separated randomly into five groups. Group I is normal mice, and groups II-V mice of generated leukemia by WEHI-3 cells. Group II-V mice were intraperitoneally injected with dimethyl sulfoxide (DMSO, as the positive control), 15, 30, and 60 mg/kg of DMC, respectively, every two days for 14 days. The body weight, blood, peritoneal fluid, liver, and spleen were individually analyzed. RESULTS: DMC did not significantly affect animal appearance and body weight. It decreased liver and spleen weight at a high dose. DMC did not affect the cluster of differentiation 3 (CD3) and CD19 cell populations but induced decrease of CD11b at 30 mg/kg treatment. However, DMC at low dose significantly increased the cluster of macrophage (Mac-3) cell populations, but at high dose it decreased them. DMC increased macrophage phagocytosis from peripheral blood mononuclear cells at 15 mg/kg treatment and peritoneal cavity at 15, 30 and 60 mg/kg of DMC treatments. DMC did not significantly affect the cytotoxic activity of natural killer (NK) cells. Furthermore, DMC decreased B and T cell proliferation at high doses. CONCLUSION: DMC elevated macrophage phagocytosis in leukemia mice in vivo.

Prenatal Diagnosis of True Fetal Mosaicism with Small Supernumerary Marker Chromosome Derived from Chromosome 16 by Funipuncture and Molecular Cytogenetics Including Chromosome Microarray.

This study examined the molecular characterization of a prenatal case with true fetal mosaicism of small supernumerary marker chromosome 16 (sSMC(16)). A 41-year-old female underwent amniocentesis at 19 weeks of gestation due to advanced maternal age. Chromosomal analysis for cultured amniocytes revealed a karyotype of 47,XY,+mar[4]/46,XY[16]. Spectral karyotyping and metaphase fluorescence in situ hybridization (FISH) demonstrated that the sSMC was derived from chromosome 16 (47,XY,+mar.ish der(16)(D16Z1+)[13/20]). Confined placental mosaicism was initially suspected because the prenatal ultrasound revealed a normal structure and the pregnancy was uneventful. However, interphase FISH of cord blood performed at 28 weeks of gestation showed 20% mosaicism of trisomy chromosome 16 (nuc ish(D16Z2×3)[40/200]). Chromosome microarray analysis further demonstrated 55% mosaicism of an 8.02 Mb segmental duplication at the subcentromeric region of 16p12.1p11.1 (arr[GRCh37] 16p12.1p11.1(27021975_35045499)×3[0.55]). The results demonstrated a true fetal mosaicism of sSMC(16) involving chromosome16p12.1p11.1 that is associated with chromosome 16p11.2 duplication syndrome (OMIM #614671). After non-directive genetic counseling, the couple opted for late termination of pregnancy. This case illustrated the use of multiple molecular cytogenetic tools to elucidate the origin and structure of sSMC, which is crucial for prenatal counseling, decision making, and clinical management.

Proteus mirabilis thoracic vertebral osteomyelitis: a case report.

BACKGROUND: Proteus mirabilis is the second most common pathogen that causes urinary tract infections after Escherichia coli. In rare cases, it is associated with vertebral osteomyelitis. The underlying mechanism of this relationship may be related to the retrograde dissemination of bacteria through the paravertebral venous plexus. CASE PRESENTATION: We report a case of an 80-year-old Taiwanese woman who had recurrent episodes of fever and chronic back pain for 1 year. All blood cultures were positive for P. mirabilis. Inflammation scans and magnetic resonance imaging revealed a previously undetected vertebral lesion between the seventh and eighth thoracic vertebra. She responded well to treatment with antibiotics, reporting considerable relief of back pain and no fever recurrence at the 4-month follow-up. CONCLUSIONS: Chronic back pain is a common but often dismissed symptom among the older population; osteomyelitis should be considered in patients with recurrent fever or neurological symptoms. Old age, chronic renal failure, and diabetes mellitus are possible predisposing factors for osteomyelitis. Our findings suggest that long-term treatment with antibiotics is effective for osteomyelitis caused by P. mirabilis,, although surgery is required for abscess formation or serious vertebral destruction.

Mangiferin induces immune responses and evaluates the survival rate in WEHI-3 cell generated mouse leukemia in vivo.

Mangiferin is a naturally occurring polyphenol, widely distributed in Thymeraceae families, and presents pharmacological activity, including anti-cancer activities in many human cancer cell lines. Mangiferin has also been reported to affect immune responses; however, no available information concerning the effects of mangiferin on immune reactions in leukemia mice in vivo. In the present study, we investigated the effects of mangiferin on leukemia WEHI-3 cell generated leukemia BLAB/c mice. Overall, the experiments were divided into two parts, one part was immune responses experiment and the other was the survival rate experiment. The immune responses and survival rate study, 40 mice for each part, were randomly separated into five groups (N = 8): Group I was normal animals and groups II-V WEHI-3 cell generated leukemia mice. Group II mice were fed normal diet as a positive control; group III, IV, and V mice received mangiferin at 40, 80, and 120 mg/kg, respectively, by intraperitoneal injection every 2 days for 20 days. Leukocytes cell population, macrophage phagocytosis, and NK cell activities were analyzed by flow cytometry. Isolated splenocytes stimulated with lipopolysaccharide (LPS) and concanavalin A (Con A) were used to determine the proliferation of B and T cells, respectively, and subsequently were analyzed by flow cytometry. Results indicated that mangiferin significantly increased body weight, decreased the liver and spleen weights of leukemia mice. Mangiferin also increased CD3 T-cell and CD19 B cell population but decreased Mac-3 macrophage and CD11b monocyte. Furthermore, mangiferin decreased phagocytosis of macrophages from PBMC and peritoneal cavity at 40, 80, and 120 mg/kg treatment. However, it also increased NK cell activity at 40 and 120 mg/kg treatment. There were no effects on T and B cell proliferation at three examined doses. In survival rate studies, mangiferin significantly elevated survival rate at 40 and 120 mg/kg treatment of leukemia mice in vivo.

Relevance of Copy Number Variation at Chromosome X in Male Fetuses Inherited from the Mother May Be Ascertained by Including Male Relatives from the Maternal Lineage in Addition to Trio Analyses.

Chromosome microarray analysis has been used for prenatal detection of copy number variations (CNVs) and genetic counseling of CNVs has been greatly improved after the accumulation of knowledge from postnatal outcomes in terms of the genotype-phenotype correlation. However, a significant number of CNVs are still regarded as variants of unknown significance (VUS). CNVs at the chromosome X (X-CNVs) represent a unique group of genetic changes in genetic counseling; X-CNVs are similar to X-linked recessive monogenic disorders in that the prognosis in males is expected to be poor. Trio analysis is typically advised to patients with X-CNVs but such an approach may be inadequate in prenatal settings since the clinical relevance is sometimes uninformative, particularly for the maternally inherited X-CNVs in male fetuses. Here, we reported four healthy women whose male fetuses were found to have X-CNVs inherited from the mothers. The X-CNVs were initially recognized as VUS or likely pathogenic in males according to the publicly available information. After extending genetic analyses to male relatives of the maternal lineages, however, the relevance of the X-CNVs was reconsidered to be likely benign. The results highlight that an extended analysis to include more relatives, in addition to the parents, provides further information for genetic counseling when X-CNVs are encountered in prenatal settings.

Ouabain induces apoptotic cell death in human prostate DU 145 cancer cells through DNA damage and TRAIL pathways.

Ouabain, a cardiotonic steroid and specific Na+ /K+ -ATPase inhibitor, has a potential to induce cancer cell apoptosis but the mechanisms of apoptosis induced by ouabain are not fully understand. The aim of this study was to investigate the cytotoxic effects of ouabain on human prostate cancer DU 145 cells in vitro. Cell morphological changes were examined by phase contrast microscopy. Cell viability, cell cycle distribution, cell apoptosis, DNA damage, the production of ROS and Ca2+ , and mitochondrial membrane potential (ΔΨm ) were measured by flow cytometry assay. Results indicated that ouabain induced cell morphological changes, decreased total cell viability, induced G0/G1 phase arrest, DNA damage, and cell apoptosis, increased ROS and Ca2+ production, but decreased the levels of ΔΨm in DU 145 cells. Ouabain also increased the activities of caspase-3, -8, and -9. Western blotting was used for measuring the alterations of apoptosis-associated protein expressions in DU 145 cells and results indicated that ouabain increased the expression of DNA damage associated proteins (pATMSer1981 , p-H2A.XSer139 , and p-p53Ser15 ) and ER-stress-associated proteins (Grp78, ATF6ß, p-PERKThr981 , PERK, eIF2A, GADD153, CaMKIIß, and caspase-4) in time-dependently. Furthermore, ouabain increased apoptosis-associated proteins (DR4, DR5, Fas, Fas Ligand, and FADD), TRAIL pathway, which related to extrinsic pathway, promoted the pro-apoptotic protein Bax, increased apoptotic-associated proteins, such as cytochrome c, AIF, Endo G, caspase-3, -8, and -9, but reduced anti-apoptotic protein Bcl-2 and Bcl-x in DU 145 cells. In conclusion, we may suggest that ouabain decreased cell viability and induced apoptotic cell death may via caspase-dependent and mitochondria-dependent pathways in human prostate cancer DU 145 cells.

Cardamonin Induces Cell Cycle Arrest, Apoptosis and Alters Apoptosis Associated Gene Expression in WEHI-3 Mouse Leukemia Cells.

Cardamonin, the chalcone class, is one of the natural components from the spicy herbaceous plant (Alpinia conchigera Griff) and has anticancer activities in many human cancer cell lines. There is, however, no information to show that cardamonin induces cell apoptosis and alters apoptosis associated gene expressions in mouse leukemia cells. Thus, we investigated the effects of cardamonin on the apoptotic cell death and associated gene expression in mouse leukemia WEHI-3 cells in vitro. Results indicated that cardamonin decreased total viable cell number via induced cell morphological changes and apoptotic cell death in WEHI-3 cells that were assay by contrast-phase microscopy and flow cytometry examinations, respectively. The flow cytometry assay indicated that cardamonin increased reactive oxygen species (ROS) and Ca 2+ production, decreased the levels of mitochondrial membrane potential ( ΔΨm) and increased caspase-3, -8 and -9 activities in WEHI-3 cells. Western blotting was performed to analyze expression of relevant pro- and anti-apoptotic proteins and results showed that cardamonin decreased anti-apoptotic protein of Bcl-2 but increased pro-apoptotic protein of Bax in WEHI-3 cells. Furthermore, cardamonin increased cytochrome c, AIF and Endo G release, increased GRP78, caspase-12 that were associated with ER stress and increased Fas, Fas-Ligand and FADD expression. Furthermore, cardamonin increased the gene expressions of DAP (death-associated protein), TMBIM4 transmembrane (BAX inhibitor motif containing 4), ATG5 (autophagy related 5) but decreased the gene expression of DDIT3 (DNA-damage inducible transcript 3), DDIT4 (DNA-damage-inducible transcript 4), BAG6 (BCL2-associated athanogene 6), BCL2L13 [BCL2-like 13 (apoptosis facilitator)] and BRAT1 (BRCA1-associated ATM activator 1) that are associated with apoptosis pathways. Based on those findings, we may suggest cardamonin induced apoptotic cell death through Fas and Fas-Ligand-, caspase- and mitochondria-dependently pathways and also affects the apoptotic gene expression in WEHI-3 cells in vitro.

Ouabain promotes immune responses in WEHI-3 cells to generate leukemia mice through enhancing phagocytosis and natural killer cell activities in vivo.

Ouabain, a cardiotonic steroid, was used for the treatment of heart failure and atrial fibrillation and induces cancer cell apoptosis in many human cancer cells including human leukemia cells. However, there are no reports to show the effects on immune responses in a leukemia mouse model. In this study, WEHI-3 cell generated leukemia mice were developed and treated by oral ouabain at 0, 0.75, 1.5, and 3 mg/kg for 15 days. Results indicated that ouabain did not affect body appearance, but decreased liver and spleen weights, B- and T-cell proliferation at all three doses treatment and increased CD19 cells at 3.0 mg/kg treatment, decreased CD3, CD11b, and Mac-3 cells levels compared with positive control. Furthermore, ouabain increased the macrophage phagocytosis from peripheral blood mononuclear cell and peritoneal cavity at all three doses treatment and increased NK cell activities. Ouabain restored GOT, GPT and LDH levels in WEHI-3 leukemia mice in vivo.

A Silicon-based Coral-like Nanostructured Microfluidics to Isolate Rare Cells in Human Circulation: Validation by SK-BR-3 Cancer Cell Line and Its Utility in Circulating Fetal Nucleated Red Blood Cells.

Circulating fetal cells (CFCs) in maternal blood are rare but have a strong potential to be the target for noninvasive prenatal diagnosis (NIPD). "Cell RevealTM system" is a silicon-based microfluidic platform capable to capture rare cell populations in human circulation. The platform is recently optimized to enhance the capture efficiency and system automation. In this study, spiking tests of SK-BR-3 breast cancer cells were used for the evaluation of capture efficiency. Then, peripheral bloods from 14 pregnant women whose fetuses have evidenced non-maternal genomic markers (e.g., de novo pathogenic copy number changes) were tested for the capture of circulating fetal nucleated red blood cells (fnRBCs). Captured cells were subjected to fluorescent in situ hybridization (FISH) on chip or recovered by an automated cell picker for molecular genetic analyses. The capture rate for the spiking tests is estimated as 88.1%. For the prenatal study, 2⁻71 fnRBCs were successfully captured from 2 mL of maternal blood in all pregnant women. The captured fnRBCs were verified to be from fetal origin. Our results demonstrated that the Cell RevealTM system has a high capture efficiency and can be used for fnRBC capture that is feasible for the genetic diagnosis of fetuses without invasive procedures.

Ursolic Acid Induces Apoptotic Cell Death Through AIF and Endo G Release Through a Mitochondria-dependent Pathway in NCI-H292 Human Lung Cancer Cells In Vitro.

BACKGROUND/AIM: Ursolic acid (UA), a triterpene compound present in natural plants, has been shown to induce cytotoxic effects on many human cancer cells through induction of cell-cycle arrest and apoptosis. This study investigated the effects of UA on human lung cancer NCI-H292 cells in vitro. MATERIALS AND METHODS: Flow cytometric assay was used to measure the percentage of cell viability, apoptotic cell death by double staining of annexin V and propidium iodide (PI), production of reactive oxygen species (ROS) and Ca2+, and mitochondriaI membrane potential (Ψm). UA-induced chromatin condensation and DNA fragmentation were examined by 4',6-diamidino-2-phenylindole staining and DNA gel electrophoresis, respectively. Western blotting was used to examine the changes of apoptosis-associated protein expression in NCI-H292 cells. RESULTS: UA reduced cell viability and induced apoptotic cell death. UA increased Ca2+ production, reduced Ψm, but did not affect ROS production in NCI-H292 cells. UA increased apoptosis-inducing factor (AIF) and endonuclease G in NCI-H292 cells. CONCLUSION: Based on these observations, we suggest UA induces apoptotic cell death via AIF and Endo G release through a mitochondria-dependent pathway in NCI-H292 cells.

Bufalin Enhances Immune Responses in Leukemic Mice Through Enhancing Phagocytosis of Macrophage In Vivo.

BACKGROUND/AIM: Bufalin, bufadienolide present in Chan Su, has been shown to induce cancer cell apoptosis in many human cancer cells, including human leukemia cells, but its effects on immune responses are unknown. MATERIALS AND METHODS: This study investigated whether bufalin affected immune responses of mice with WEHI-3 cell-generated leukemia in vivo. BALB/c mice were intraperitoneally injected with WEHI-3 cells to develop leukemia and then were treated with oral treatment with bufalin at different doses (0, 0.1, 0.2 and 0.4 mg/kg) for 2 weeks. At the end of treatment, all mice were weighted and blood was collected; liver and spleen tissues were collected for cell marker, phagocytosis, natural killer (NK) cell activity and T- and B-cell proliferation measurements by using flow cytometric assays. RESULTS: When compared with the leukemia control group, bufalin increased the body weight, but reduced liver and spleen weights, and reduced CD3, CD16 and Mac-3 cell markers at 0.4 mg/kg treatment and increased CD11b marker at 0.1 and 0.2 mg/kg treatment. Furthermore, bufalin at 0.4 mg/kg increased phagocytosis by macrophages isolated from peripheral blood mononuclear cells and at 0.1 mg/kg by those from the peritoneal cavity. Bufalin (0.2 and 0.4 mg/kg) increased NK cell cytotoxic activity at effector:target ratio of 50:1. Bufalin increased B-cell proliferation at 0.1 and 0.2 mg/kg treatment but only increased T-cell proliferation at 0.1 mg/kg. Bufalin increased glutamate oxaloacetate transaminase level at all dose treatments, increased glutamic pyruvic transaminase level only at 0.1 mg/kg treatment, but reduced the level of lactate dehydrogenase at all dose levels in mice with WEHI-3 cell-induced leukemia in vivo. CONCLUSION: Bufalin increased immune responses by enhancing phagocytosis in mice with leukemia mice.