This study aimed to accurately analyze the relationship between calcium oxalate (CaOx) stone formation and the components of urinary nanocrystallites.
High-resolution transmission electron microscopy (HRTEM), selected area electron diffraction, fast Fourier transformation of HRTEM, and energy dispersive X-ray spectroscopy were performed to analyze the components of these nanocrystallites.
The main components of CaOx stones are calcium oxalate monohydrate and a small amount of dehydrate, while those of urinary nanocrystallites are calcium oxalate monohydrate, uric acid, and calcium phosphate. The mechanism of formation of CaOx stones was discussed based on the components of urinary nanocrystallites.
The formation of CaOx stones is closely related both to the properties of urinary nanocrystallites and to the urinary components. The combination of HRTEM, fast Fourier transformation, selected area electron diffraction, and energy dispersive X-ray spectroscopy could be accurately performed to analyze the components of single urinary nanocrystallites. This result provides evidence for nanouric acid and/or nanocalcium phosphate crystallites as the central nidus to induce CaOx stone formation.
nanocrystallites; calcium oxalate stones; heterogeneous nucleation; HRTEM; EDS
Bone marrow transplantation might be an effective method to cure type 1 diabetes mellitus. This study aimed to investigate whether bone marrow transplantation could reverse hyperglycemia in diabetic mice and whether high-dose total body irradiation followed by high-dose bone marrow mononuclear cell infusion could improve the efficiency of bone marrow transplantation in treating diabetic mice. Diabetic mice after multiple low doses of streptozotocin injection were irradiated followed by infusion with approximately 1×107 bone marrow mononuclear cells intravenously. Before and after bone marrow transplantation, fasting blood glucose, intraperitoneal glucose tolerance test, serum insulin, pancreatic histology, and the examination of insulin and glucagon in islets were processed. All recipients returned to near euglycemic within 1 week after undergoing bone marrow transplantation. No mice became hyperglycemia again during investigation period. The change of serum insulin, glucose tolerance test, pancreatic histology and the expression of insulin and glucagon in recipient islets after bone marrow transplantation all revealed islets regeneration and significant amelioration when compared respectively with those of diabetic mice without bone marrow transplantation. Bone marrow transplantation contributed to reduce blood glucose, prevent further blood glucose hike in diabetic recipients, and promote islets regeneration. High-dose total body irradiation in combination with high-dose bone marrow monoclear cell infusion could improve the efficiency of bone marrow transplantation in treating streptozotocin-induced diabetes.
Type 1 diabetes; bone marrow transplantation; streptozotocin; total body irradiation
Aggressive natural killer cell leukemia (ANKL) is a rare hematological malignancy that is particularly common among the Asian population. In the current study, we retrospectively evaluated six Chinese ANKL patients, including five males and one female, with a median age of 42 years (range 22 to 50 years). A number of unusual pathogenic manifestations were found in these ANKL patients, such as isolated extraocular muscle involvement, and hemophagocytic syndrome (HPS) with acute renal failure and multiple cavity effusion. Four of the patients died between two and six months after the diagnosis; however, there were two ANKL cases whose clinical behavior differed from the typical clinical course. One survived for over 30 months after splenectomy and chemotherapy treatment, and another ANKL case derived from chronic lymphoproliferative disorders of NK-cells (CLPD-NK) was treated with allogeneic bone marrow transplant (allo-BMT) and survived over 18 months. In conclusion, four cases experienced an aggressive clinical course whereas two demonstrated an indolent manifestation of their disease. New therapeutic regimens including allo-BMT should be optimized in order to improve outcomes of this disease.
Aggressive NK-cell leukemia; lymphoproliferative disorders of NK-cells; hemophagocytic syndrome; retrospective study
Celastrol is an active compound extracted from the root bark of the traditional Chinese medicine Tripterygium wilfordii Hook F. To investigate the effect of celastrol on human multiple myeloma cell cycle arrest and apoptosis and explore its molecular mechanism of action. The activity of celastrol on LP-1 cell proliferation was detected by WST-8 assay. The celastrol-induced cell cycle arrest was analyzed by flow cytometry after propidium iodide staining. Nuclear translocation of the nuclear factor kappa B (NF-κB) was observed by fluorescence microscope. Celastrol inhibited cell proliferation of LP-1 myeloma cell in a dose-dependent manner with IC50 values of 0.8817 µM, which was mediated through G1 cell cycle arrest and p27 induction. Celastrol induced apoptosis in LP-1 and RPMI 8226 myeloma cells in a time and dose dependent manner, and it involved Caspase-3 activation and NF-κB pathway. Celastrol down-modulated antiapoptotic proteins including Bcl-2 and survivin expression. The expression of NF-κB and IKKa were decreased after celastrol treatment. Celastrol effectively blocked the nuclear translocation of the p65 subunit and induced human multiple myeloma cell cycle arrest and apoptosis by p27 upregulation and NF-kB modulation. It has been demonstrated that the effect of celastrol on NF-kB was HO-1-independent by using zinc protoporphyrin-9 (ZnPPIX), a selective heme oxygenase inhibitor. From the results, it could be inferred that celastrol may be used as a NF-kB inhibitor to inhibit myeloma cell proliferation.
Diethyl citrate (Et2Cit) is a new potential anticoagulant. The coordination dynamics and coordination mechanism of Et2Cit with Ca2+ ions and the effect of pH on the complex were examined. The result was compared with that for the conventional anticoagulant sodium citrate (Na3Cit). The reaction order (n) of Et2Cit and Na3Cit with Ca2+ was 2.46 and 2.44, respectively. The reaction rate constant (k) was 120 and 289 L·mol−1·s−1. The reverse reaction rate constant (kre) was 0.52 and 0.15 L·mol−1·s−1, respectively. It is indicated that the coordination ability of Et2Cit with Ca2+ was weaker than that of Na3Cit. However, the dissociation rate of the calcium complex of Et2Cit was faster than that of Na3Cit. Increased pH accelerated the dissociation rate of the complex and improved its anticoagulant effect. The Et2Cit complex with calcium was synthesized and characterized by elemental analysis, XRD, FT-IR, 1H NMR, and ICP. These characteristics indicated that O in –COOH and C–O–C of Et2Cit was coordinated with Ca2+ in a bidentate manner with 1 : 1 coordination proportion; that is, complex CaEt2Cit was formed. Given that CaEt2Cit released Ca2+ more easily than Na3Cit, a calcium solution was not needed in intravenous infusions using Et2Cit as anticoagulant unlike using Na3Cit. Consequently, hypocalcemia and hypercalcemia were avoided.
The influences of chondroitin sulfate C (C6S) on size, aggregation, sedimentation, and Zeta potential of sub-micron calcium oxalate monohydrate (COM) and calcium oxalate dihydrate (COD) crystallites with mean sizes of about 330 nm were investigated using an X-ray diffractometer, nanoparticle size Zeta potential analyzer, ultraviolet spectrophotometer, and scanning electron microscope, after which the results were compared with those of micron-grade crystals. C6S inhibited the conversion of COD to COM and the aggregation of COM and COD crystallitesis; it also decreased their sedimentation rate, thus increasing their stability in aqueous solution. The smaller the size of the COD crystallites, the easier they can be converted to COM. The stability of sub-micron COD was worse than that of micron-grade crystals. C6S can inhibit the formation of calcium oxalate stones.
The changes in urinary crystal properties in patients with calcium oxalate (CaOx) calculi after oral administration of potassium citrate (K3cit) were investigated via atomic force microscopy (AFM), scanning electron microscopy (SEM), X-ray powder diffractometry (XRD), and zeta potential analyzer. The AFM and SEM results showed that the surface of urinary crystals became concave, the edges and corners of crystals became blunt, the average size of urinary crystallines decreased significantly, and aggregation of urinary crystals was reduced. These changes were attributed to the significant increase in concentration of excreted citrate to 492 ± 118 mg/L after K3cit intake from 289 ± 83 mg/L before K3cit intake. After the amount of urinary citrate was increased, it complexed with Ca2+ ions on urinary crystals, which dissolved these crystals. Thus, the appearance of concave urinary crystals was a direct evidence of CaOx dissolution by citrate in vivo. The XRD results showed that the quantities and species of urinary crystals decreased after K3cit intake. The mechanism of inhibition of formation of CaOx stones by K3cit was possibly due to the complexation of Ca2+ with citrate, increase in urine pH, concentration of urinary inhibitor glycosaminoglycans (GAGs), and the absolute value of zeta potential after K3cit intake.
Like that of many protein-coding genes, expression of the p21CIP1 cell cycle inhibitor is regulated at the level of transcription elongation. While many transcriptional activators have been shown to stimulate elongation, the mechanisms by which promoter-specific repressors regulate pausing and elongation by RNA polymerase II (RNA PolII) are not well described. Here we report that the transcription factor Sp3 inhibits basal p21CIP1 gene expression by promoter-bound RNA PolII. Knockdown of Sp3 led to increased p21CIP1 mRNA levels and reduced occupancy of the negative elongation factor (NELF) at the p21CIP1 promoter, although the level of binding of the positive transcription elongation factor b (P-TEFb) kinase was not increased. Sp3 depletion correlated with increased H3K36me3 and H2Bub1, two histone modifications associated with transcription elongation. Further, Sp3 was shown to promote the binding of protein phosphatase 1 (PP1) to the p21CIP1 promoter, leading to reduced H3S10 phosphorylation, a finding consistent with Sp3-dependent regulation of the local balance between kinase and phosphatase activities. Analysis of other targets of Sp3-mediated repression suggests that, in addition to previously described SUMO modification-dependent chromatin-silencing mechanisms, inhibition of the transition of paused RNA PolII to productive elongation, described here for p21CIP1, is a general mechanism by which transcription factor Sp3 fine-tunes gene expression.
To determine whether the use of idarubicin+cytarabine (IA) is more effective than the use of daunorubicin+cytarabine (DA) as induction chemotherapy for patients with newly diagnosed acute myeloid leukaemia.
A computer-based search was performed. Randomised trials comparing IA with DA as induction therapy for newly diagnosed AML were included in this meta-analysis. The primary outcome of interest for our analysis was survival (disease-free survival, event-free survival and overall survival); the secondary endpoint was complete remission.
Ten trials with 4,060 patients were eligible for this meta-analysis. Our pooled results suggest that IA is associated with a significant advantage in CR (RR = 1·23; 95% CI = 1·07–1·41, p = 0.004), EFS (HR = 0·64; 95% CI = 0·45–0·91, p = 0.013), and OS (HR = 0·88; 95% CI = 0·81–0·95, p = 0.02) but not in DFS (HR = 0·90; 95% CI = 0·80–1·00, p = 0.06). In the subgroup analysis, age had a significant interaction with OS and CR benefits.
Our analysis indicated that IA could improve the duration of overall survival compared to DA as induction therapy for young patients with newly diagnosed AML. Further study is needed to determine whether IA can produce clinical benefits in selected genetic or molecular subgroups of young AML patients.
To study the mechanisms in gambogic acid (GA) -induced JeKo-1 human Mantle Cell Lymphoma cell apoptosis in vitro.
The proliferation of GA-treated JeKo-1 cells was measured by CCK-8 assay and Ki-67 immunocytochemical detection. Apoptosis, cell cycle and mitochondrial membrane potential were measured by flow cytometric analysis. Caspase-3, -8 and -9 were detected by colorimetric assay. Bcl-2 and Bax were analyzed by Western blotting.
GA inhibited cell growth in a time- and dose- dependent manner. GA induces apoptosis in JeKo-1 cells but not in normal bone marrow cells, which was involved in reducing the membrane potential of mitochondria, activating caspases-3, -8 and -9 and decreasing the ratio of Bcl-2 and Bax without cell cycle arresting.
GA induced apoptosis in human MCL JeKo-1 cells by regulating Bcl-2/Bax and activating caspase-3, -8 and -9 via mitochondrial pathway without affecting cell cycle.
Gambogic acid; JeKo-1 cells; cell cycle arrest, apoptosis; membrane potential of mitochondria; caspase-3; caspase-8; caspase-9; Bax; Bcl-2
The property changes of urinary nanocrystallites in 13 patients with calcium oxalate (CaOx) stones were studied before and after ingestion of potassium citrate (K3cit), a therapeutic drug for stones. The analytical techniques included nanoparticle size analysis, transmission electron microscopy, X-ray diffraction, and Fourier-transform infrared spectroscopy. The studied properties included the components, morphologies, zeta potentials, particle size distributions, light intensity autocorrelation curves, and polydispersity indices (PDIs) of the nanocrystallites. The main components of the urinary nanocrystallites before K3cit intake included uric acid, β-calcium phosphate, and calcium oxalate monohydrate. After K3cit intake, the quantities, species, and percentages of aggregated crystals decreased, whereas the percentages of monosodium urate and calcium oxalate dehydrate increased, and some crystallites became blunt. Moreover, the urinary pH increased from 5.96 ± 0.43 to 6.46 ± 0.50, the crystallite size decreased from 524 ± 320 nm to 354 ± 173 nm, and the zeta potential decreased from −4.85 ± 2.87 mV to −8.77 ± 3.03 mV. The autocorrelation curves became smooth, the decay time decreased from 11.4 ± 3.2 ms to 4.3 ± 1.7 ms, and the PDI decreased from 0.67 ± 0.14 to 0.53 ± 0.19. These changes helped inhibit CaOx calculus formation.
nanotechnology; zeta potential; aggregation
This study aims to investigate the adhesion characteristics between submicron calcium oxalate dihydrate (COD) with a size of 150 ± 50 nm and African green monkey kidney epithelial cells (Vero cells) before and after damage, and to discuss the mechanism of kidney stone formation.
Vero cells were oxidatively injured by hydrogen peroxide to establish a model of injured cells. Scanning electron microscopy was used to observe Vero–COD adhesion. Inductively coupled plasma emission spectrometry was used to quantitatively measure the amount of adhered COD microcrystals. Nanoparticle size analyzer and laser scanning confocal microscopy were performed to measure the change in the zeta potential on the Vero cell surface and the change in osteopontin expression during the adhesion process, respectively. The level of cell injury was evaluated by measuring the changes in malonaldehyde content, and cell viability during the adhesion process.
The adhesion capacity of Vero cells in the injury group to COD microcrystals was obviously stronger than that of Vero cells in the control group. After adhesion to COD, cell viability dropped, both malonaldehyde content and cell surface zeta potential increased, and the fluorescence intensity of osteopontin decreased because the osteopontin molecules were successfully covered by COD. Submicron COD further damaged the cells during the adhesion process, especially for Vero cells in the control group, leading to an elevated amount of attached microcrystals.
Submicron COD can further damage injured Vero cells during the adhesion process. The amount of attached microcrystals is proportional to the degree of cell damage. The increased amount of microcrystals that adhered to the injured epithelial cells plays an important role in the formation of early-stage kidney stones.
cell modulation; crystal adhesion; calcium oxalate dihydrate; kidney stone; pathological mineralization
Epigenetic regulation of gene expression by histone-modifying co-repressor complexes is central to normal animal development. The NAD+-dependent deacetylase and gene repressor SIRT1 removes histone H4K16 acetylation marks and facilitates heterochromatin formation. However, the mechanistic contribution of SIRT1 to epigenetic regulation at euchromatic loci and whether it acts in concert with other chromatin-modifying activities to control developmental gene expression programs remain unclear. We describe here a SIRT1 co-repressor complex containing the histone H3K4 demethylase LSD1/KDM1A and several other LSD1-associated proteins. SIRT1 and LSD1 interact directly and play conserved and concerted roles in H4K16 deacetylation and H3K4 demethylation to repress genes regulated by the Notch signaling pathway. Mutations in Drosophila SIRT1 and LSD1 orthologs result in similar developmental phenotypes and genetically interact with the Notch pathway in Drosophila. These findings offer new insights into conserved mechanisms of epigenetic gene repression and regulation of development by SIRT1 in metazoans.
SIRT1; LSD1; co-repressor; Notch; chromatin modification
Background and Objectives
Several trials have generated conflicting results about the results of high-dose chemotherapy followed by autologous stem cell transplantation (HDCT) for primary breast cancer. This meta-analysis summarizes the available evidence from all suitable studies.
Design and Methods
Prospective, randomized trials with HDCT as a first-line therapy for primary breast cancer were included in this meta-analysis. The primary outcome of interest for our analysis was survival (disease-free survival and overall survival); secondary endpoints included treatment-related mortality (TRM) and second (non-breast) cancers. We used a median age of 47, a PR positive rate of 50% and a premenopausal rate of 70% as cutoff values to complete the subgroup analyses, which were pre-planned according to the prepared protocol.
Fourteen trials with 5747 patients were eligible for the meta-analysis. Compared with non-HDCT, non-significant second (non-breast) cancers (RR = 1.28; 95% CI = 0.82–1.98) and higher TRM (RR = 3.42; 95% CI = 1.32–8.86) were associated with HDCT for primary breast cancer. A significant DFS benefit of HDCT was documented (HR = 0.89; 95% CI = 0.79–0.99). No difference in OS (overall survival) was found when the studies were pooled (HR = 0.91; 95% CI = 0.82–1.00, p = 0.062). In subgroup analysis, age and hormone receptor status had a significant interaction with prolonged DFS and OS.
HDCT has a benefit on DFS and OS compared to SDC in some special patients with high-risk primary breast cancer.
Post-translational modification of transcription factors by the small ubiquitin related modifier SUMO is associated with transcriptional repression, but the underlying mechanisms remain incompletely described. We have identified binding of the LSD1/CoREST1/HDAC co-repressor complex to SUMO-2. Here we show that CoREST1 binds directly and non-covalently to SUMO-2, but not SUMO-1, and CoREST1 bridges binding of the histone demethylase LSD1 to SUMO-2. Depletion of SUMO-2/3 conjugates led to transcriptional de-repression, reduced occupancy of CoREST1 and LSD1 and changes in histone methylation and acetylation at some, but not all, LSD1/CoREST1/HDAC target genes. We have identified a non-consensus SUMO-interaction motif (SIM) in CoREST1 required for SUMO-2 binding and we show that mutation of the CoREST1 SIM disrupted SUMO-2 binding and transcriptional repression of some neuronal specific genes in non-neuronal cells. Our results reveal that direct interactions between CoREST1 and SUMO-2 mediate SUMO-dependent changes in chromatin structure and transcription important for cell type-specific gene expression.
The composition and morphology of nanocrystals in urines of healthy persons and lithogenic patients were comparatively investigated by means of X-ray diffraction (XRD) and transmission electron microscopy (TEM). It was shown that the main composition of urinary nanocrystals in healthy persons were calcium oxalate dihydrate (COD), uric acid, and ammonium magnesium phosphate (struvite). However, the main compositions of urinary nanocrystals in lithogenic patients were struvite, β-tricalcium phosphate, uric acid, COD, and calcium oxalate monohydrate (COM). According to the XRD data, the size of nanocrystals was calculated to be 23∼72 nm in healthy urine and 12∼118 nm in lithogenic urine by Scherer formula. TEM results showed that the nanocrystals in healthy urine were dispersive and uniform with a mean size of about 38 nm. In contrast, the nanocrystals in lithogenic urine were much aggregated with a mean size of about 55 nm. The results in this work indicated that the urinary stone formation may be prevented by diminishing the aggregation and the size differentiation of urinary nanocrystals by physical or chemical methods.
Posttranslational modification of substrates by the small ubiquitin-like modifier, SUMO, regulates diverse biological processes, including transcription, DNA repair, nucleocytoplasmic trafficking, and chromosome segregation. SUMOylation is reversible, and several mammalian homologs of the yeast SUMO-specific protease Ulp1, termed SENPs, have been identified. We demonstrate here that SENP5, a previously uncharacterized Ulp1 homolog, has SUMO C-terminal hydrolase and SUMO isopeptidase activities. In contrast to other SENPs, the C-terminal catalytic domain of SENP5 preferentially processed SUMO-3 compared to SUMO-1 precursors and preferentially removed SUMO-2 and SUMO-3 from SUMO-modified RanGAP1 in vitro. In cotransfection assays, SENP5 preferentially reduced high-molecular-weight conjugates of SUMO-2 compared to SUMO-1 in vivo. Full-length SENP5 localized to the nucleolus. Deletion of the noncatalytic N-terminal domain led to loss of nucleolar localization and increased de-SUMOylation activity in vivo. Knockdown of SENP5 by RNA interference resulted in increased levels of SUMO-1 and SUMO-2/3 conjugates, inhibition of cell proliferation, defects in nuclear morphology, and appearance of binucleate cells, revealing an essential role for SENP5 in mitosis and/or cytokinesis. These findings establish SENP5 as a SUMO-specific protease required for cell division and suggest that mechanisms involving both the catalytic and noncatalytic domains determine the distinct substrate specificities of the mammalian SUMO-specific proteases.