PE-Cyanine7 anti-CD19, Cell Cycle Analysis (PI / RNASE Solution) and FITC Annexin V Apoptosis Detection Kit with PI.

An article published this year in “JOURNAL OF HEMATOLOGY & ONCOLOGY” using our PE-Cyanine7 anti-CD19, Cell Cycle Analysis (PI / RNASE Solutiuon) and FITC Annexin V Apoptosis Detection Kit with PI, by our customers from Cancer Research Center-IBMCC (USAL-CSIC), Department of Hematology, University Hospital of Salamanca, Institute of Biomedical Research of Salamanca (IBSAL), Salamanca, Spain, in the analysis of how DEPTOR maintains plasma cell differentiation and favorably affects prognosis in multiple myeloma. Congrats and Thanks.

Summary:

Background

The B cell maturation process involves multiple steps, which are controlled by relevant pathways and transcription factors. The understanding of the final stages of plasma cell (PC) differentiation could provide new insights for therapeutic strategies in multiple myeloma (MM). Here, we explore the role of DEPTOR, an mTOR inhibitor, in the terminal differentiation of myeloma cells, and its potential impact on patient survival.

Methods

The expression level of DEPTOR in MM cell lines and B cell populations was measured by real-time RT-PCR, and/or Western blot analysis. DEPTOR protein level in MM patients was quantified by capillary electrophoresis immunoassay. RNA interference was used to downregulate DEPTOR in MM cell lines.

Results

DEPTOR knockdown in H929 and MM1S cell lines induced dedifferentiation of myeloma cells, as demonstrated by the upregulation of PAX5 and BCL6, the downregulation of IRF4, and a clear reduction in cell size and endoplasmic reticulum mass. This effect seemed to be independent of mTOR signaling, since mTOR substrates were not affected by DEPTOR knockdown. Additionally, the potential for DEPTOR to be deregulated in MM by particular miRNAs was investigated. The ectopic expression of miR-135b and miR-642a in myeloma cell lines substantially diminished DEPTOR protein levels, and caused dedifferentiation of myeloma cells. Interestingly, the level of expression of DEPTOR protein in myeloma patients was highly variable, the highest levels being associated with longer progression-free survival.

Conclusions

Our results demonstrate for the first time that DEPTOR expression is required to maintain myeloma cell differentiation and that high level of its expression are associated with better outcome.

Reference: https://jhoonline.biomedcentral.com/articles/10.1186/s13045-017-0461-8

Product link:                      

PE-Cyanine7 - anti Human CD19 - HIB19: http://www.immunostep.com/cd-antibodies/2765-cd19.html

Cell Cycle Analysis (PI / RNASE Solutiuon): http://www.immunostep.com/solutions-buffers-chemicals/1289-pi-rnase-solution.html

FITC Annexin V Apoptosis Detection Kit with PI: http://www.immunostep.com/apoptosis-tools/3727-anxvkf-100t.html

PE-Cyanine7 - anti Human CD19 - HIB19

An article published this year in “JOURNAL OF HEMATOLOGY & ONCOLOGY” using our PE-Cyanine7 anti-CD19, by our customers from Servicio de Hematología & IBSAL, IBMCC, CIC Universidad de Salamanca-CSIC, Hospital Universitario, in the analysis of how Next-generation sequencing and FISH studies reveal the appearance of gene mutations and chromosomal abnormalities in hematopoietic progenitors in chronic lymphocytic leukemia. Congrats and Thanks.

Summary:

Background

Chronic lymphocytic leukemia (CLL) is a highly genetically heterogeneous disease. Although CLL has been traditionally considered as a mature B cell leukemia, few independent studies have shown that the genetic alterations may appear in CD34+ hematopoietic progenitors. However, the presence of both chromosomal aberrations and gene mutations in CD34+ cells from the same patients has not been explored.

Methods

Amplicon-based deep next-generation sequencing (NGS) studies were carried out in magnetically activated-cell-sorting separated CD19+ mature B lymphocytes and CD34+ hematopoietic progenitors (n = 56) to study the mutational status of TP53, NOTCH1, SF3B1, FBXW7, MYD88, and XPO1 genes. In addition, ultra-deep NGS was performed in a subset of seven patients to determine the presence of mutations in flow-sorted CD34+CD19− early hematopoietic progenitors. Fluorescence in situ hybridization (FISH) studies were performed in the CD34+ cells from nine patients of the cohort to examine the presence of cytogenetic abnormalities.

Results

NGS studies revealed a total of 28 mutations in 24 CLL patients. Interestingly, 15 of them also showed the same mutations in their corresponding whole population of CD34+ progenitors. The majority of NOTCH1 (7/9) and XPO1 (4/4) mutations presented a similar mutational burden in both cell fractions; by contrast, mutations of TP53 (2/2), FBXW7 (2/2), and SF3B1 (3/4) showed lower mutational allele frequencies, or even none, in the CD34+ cells compared with the CD19+ population. Ultra-deep NGS confirmed the presence of FBXW7, MYD88, NOTCH1, and XPO1 mutations in the subpopulation of CD34+CD19− early hematopoietic progenitors (6/7). Furthermore, FISH studies showed the presence of 11q and 13q deletions (2/2 and 3/5, respectively) in CD34+ progenitors but the absence of IGH cytogenetic alterations (0/2) in the CD34+ cells. Combining all the results from NGS and FISH, a model of the appearance and expansion of genetic alterations in CLL was derived, suggesting that most of the genetic events appear on the hematopoietic progenitors, although these mutations could induce the beginning of tumoral cell expansion at different stage of B cell differentiation.

Conclusions

Our study showed the presence of both gene mutations and chromosomal abnormalities in early hematopoietic progenitor cells from CLL patients.

Reference:

https://jhoonline.biomedcentral.com/articles/10.1186/s13045-017-0450-y

Product link:

PE-Cyanine7 - anti Human CD19 - HIB19: http://www.immunostep.com/cd-antibodies/2765-cd19.html

FITC Annexin V Apoptosis Detection Kit with 7-AAD.

An article published this year in “NUCLEIC ACIDS RESEARCH” using our Annexin V-FITC and 7-aminoactinomycin D (7-AAD), by our customers from Hospital Universitario La Paz-IdiPAZ,  and CIBER de Bioingenieria, Biomateriales y Nanomedicina, CIBER-BBN, Madrid, Spain, in the study of New inhibitor targeting human transcription factor HSF1: effects on the heat shock response and tumor cell survival. Congrats and Thanks.

Summary:

Comparative modeling of the DNA-binding domain of human HSF1 facilitated the prediction of possible binding pockets for small molecules and definition of corresponding pharmacophores. In silico screening of a large library of lead-like compounds identified a set of compounds that satisfied the pharmacophoric criteria, a selection of which compounds was purchased to populate a biased sublibrary. A discriminating cell-based screening assay identified compound 001, which was subjected to systematic analysis of structure–activity relationships, resulting in the development of compound 115 (IHSF115). IHSF115 bound to an isolated HSF1 DNA binding domain fragment. The compound did not affect heat-induced oligomerization, nuclear localization and specific DNA binding but inhibited the transcriptional activity of human HSF1, interfering with the assembly of ATF1-containing transcription complexes. IHSF115 was employed to probe the human heat shock response at the transcriptome level. In contrast to earlier studies of differential regulation in HSF1-naıve and -depleted cells, our results suggest that a large majority of heat-induced genes is positively regulated by HSF1. That IHSF115 effectively countermanded repression in a significant fraction of heat-repressed genes suggests that repression of these genes is mediated by transcriptionally active

HSF1. IHSF115 is cytotoxic for a variety of human cancer cell lines, multiple myeloma lines consistently exhibiting high sensitivity.

Reference:

http://vuh-la-uhra.herts.ac.uk/bitstream/handle/2299/17806/gkx194.pdf?sequence=2

Product link:

FITC Annexin V Apoptosis Detection Kit with 7-AAD: http://www.immunostep.com/apoptosis-tools/3741-anxvkf-100t.html

CD105‑FITC

An article published this year in “EXPERIMENTAL AND THERAPEUTIC MEDICINE” using our CD105‑FITC, by our customers from Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brasil, in the study of how Hair follicle-derived mesenchymal cells support undifferentiated growth of embryonic stem cells. Congrats and Thanks.

Summary:

The aim of the present study was to investigate whether feeder layers composed of human hair follicle-derived mesenchymal stem cells (hHFDCs) are able to support human embryonic stem cells (hESCs). hHFDCs and mouse embryonic fibroblasts (MEFs) were isolated and cultured in Dulbecco's modified Eagle's medium (DMEM)/F‑12 and low‑glucose DMEM, respectively. hHFDCs were passaged three times and subsequently characterized. hHFDCs and MEFs were mitotically inactivated with mitomycin C for 3 h prior to co‑culture with H9‑hESCs. hESCs were initially established on a mouse feeder layer, subsequently transferred onto a human feeder layer and split every 5 days. Cell morphology, expression of specific ‘undifferentiation’ markers and growth factors, and the differentiation capacity of hESCs grown on the hHFDC feeder layer were analyzed. hHFDCs are adherent to plastic, possess the classic mesenchymal stem cell phenotype [they express cluster of differentiation (CD)90, CD73 and CD105] and are able to differentiate into adipocytes, chondroblasts and osteocytes, indicating that these cells are multipotent. Population‑doubling time analysis revealed that hHFDCs rapidly proliferate over 34.5 h. As a feeder layer, hHFDC behaved similarly to MEF in maintaining the morphology of hESCs. The results of alkaline phosphatase activity, reverse transcription‑quantitative polymerase chain reaction analysis of the expression of pluripotency transcription factors [octamer‑binding transcription factor 4 (Oct4), Nanog and sex determining region Y‑box 2], and immunofluorescence assays of markers (stage‑specific embryonic antigen‑4 and Oct4) in hESCs co‑cultured over hHFDC, indicated that the undifferentiated state of hESCs was preserved. No change in the level of growth factor transcripts (bone morphogenetic protein 4, fibroblast growth factor‑2, vascular endothelial growth factor, Pigment epithelium‑derived factor and transforming growth factor‑β1) was detected for either feeder layer prior to or following inactivation. Similar phenotypes of embryoid body formation, size and morphology were observed in the hHFDC and MEF feeders. In conclusion, hHFDC maintained hESCs in an undifferentiated state comparable to MEF in standard conditions, which may be an important finding regarding the establishment of stem cell-based translational applications.

Reference:

https://www.spandidos-publications.com/10.3892/etm.2017.4195

Product link:

CD105 FITC: http://www.immunostep.com/antibodies/2792-cd105.html

Annexin V-FITC Apoptosis Detection Kit

An article published this year in “CELLULAR PHYSIOLOGY AND BIOCHEMISTRY” using our Annexin V-FITC Apoptosis Detection Kit, by our customers from Institute for Maternal and Child Health – IRCCS “Burlo Garofolo” – Trieste, University of Trieste, Trieste, Italy, in the study of Lack of Prenylated Proteins, Autophagy Impairment and Apoptosis in SH-SY5Y Neuronal Cell Model of Mevalonate Kinase Deficiency. Congrats and Thanks.

Summary:

Background/Aims: Mevalonate Kinase Deficiency (MKD), is a hereditary disease due to mutations in mevalonate kinase gene (MVK). MKD has heterogeneous clinical phenotypes: the correlation between MVK mutations and MKD clinical phenotype is still to be fully elucidated. Deficiency of prenylated proteins has been hypothesized as possible MKD pathogenic mechanism. Based on this hypothesis and considering that neurologic impairment characterizes Mevalonic Aciduria (MA), the most severe form of MKD, we studied the effects of I268T and N301T MVK mutations on protein prenylation, autophagy and programmed cell death in SH-SY5Y neuroblastoma cell lines. Methods: SH-SY5Y cells were transiently transfected, with the pCMV-6 plasmid containing MVK wild type and the two mutated sequences. Protein prenylation levels were evaluated using GFP-RhoA-F to assess farnesylation, and GFP-RhoA to evaluate geranylgeranylation; autophagy was measured by evaluating LC3 and p62 protein levels, while Annexin V-FITC and Propidium Iodide staining allowed apoptosis detection. Results: MVK mutants’ over-expression causes decreased levels of farnesylation and geranylgeranylation, and also increased LC3 lipidation in SH-SY5Y, with concomitant p62 accumulation. Treatment with bafilomycin A1 (an inhibitor of vacuolar H+-ATPase, a late autophagy inhibitor) further increase LC3-II and p62 levels, suggesting that degradation of autophagolysosome could be impaired. SH-SY5Y, with both MVK mutants, showed apoptosis increase; the presence of N301T associated with augmented cell death. Conclusions: We hypothesize that mevalonate pathway impairment causes alteration of farnesylation and geranylgeranylation proteins and alteration of the autophagic flux; these changes can induce apoptosis, possibly more relevant in the presence of N301T mutation.

Reference:

https://www.karger.com/Article/FullText/471235

Product link:

FITC Annexin V Apoptosis Detection Kit: http://www.immunostep.com/apoptosis-tools/3741-anxvkf-100t.html

Annexin V-FITC Apoptosis Detection Kit.

An article published this year in “JOURNAL OF MATERIALS CHEMISTRY B” using our Annexin V-FITC Apoptosis Detection Kit, by our customers from Institut Lavoisier, UMR 8180 CNRS Universite´ de Versailles Saint-Quentin-en-Yvelines, Versailles, France, in the study of how Crystal structure dependent in vitro antioxidant activity of biocompatible calcium gallate MOFs. Congrats and Thanks.

Summary:

Two novel 3-D coordination polymers, denoted MIL-155 and MIL-156 (MIL stands for Materials Institute Lavoisier), built up from calcium and the naturally occurring gallic acid (H4gal), have been hydrothermally synthesized and their crystal structures were determined by single-crystal X-ray diffraction. These solids are based on different inorganic subunits: infinite chains of edge-sharing dimers of CaO7 polyhedra linked through partially deprotonated gallate ligands (H2gal2_) for MIL-155 or [Ca2(H2O)(H2gal)2]_2H2O, and ribbon-like inorganic subunits containing both eight-fold or six-fold coordinated CaII ions linked through fully deprotonated gallate ligands (gal4_) for MIL-156 or [Ca3K2(H2O)2(gal)2]_nH2O (n B 5). Both solids contain small channels filled with water molecules, with, however no accessible porosity towards N2 at 77 K. MIL-155 and MIL-156 were proven to be biocompatible, as evidenced by in vitro assays (viability and cell proliferation/death balance). While the high chemical stability of MIL-156 makes it almost bioinert, the progressive degradation of MIL-155 leads to an important protective antioxidant effect, associated with the release of the bioactive gallate ligand.

Reference:

http://pubs.rsc.org/en/content/articlehtml/2017/tb/c6tb03101c

Product link:

FITC Annexin V Apoptosis Detection Kit: http://www.immunostep.com/apoptosis-tools/3741-anxvkf-100t.html