Ohbuchi A, et al. Blood Res. 2018 Jun;53(2):177-180. https://doi.org/10.5045/br.2018.53.2.177

Acute promyelocytic leukemia (APL) is characterized by the specific PML-RARa fusion gene resulting from translocation t(15;17) (q22;q12). Internal tandem duplication (ITD) of the FLT3 gene has been observed in approximately 35% of APLs, and large-scale studies have identified the presence of ITD as an adverse prognostic factor for acute myeloblastic leukemia (AML) patients. Aberrant expressions of surface antigens, such as CD2, CD34, and CD56, have been found in APL, but the implications of this are not well understood. We investigated the incidence of the FLT3/ITD mutation and FLT3/D835 (I836) point mutation in 25 APL patients. Incidence ratios of FLT3/ITD, D835 (I836), and both FLT3/ITD and D835 (I836) were 36%, 36% and 8%, respectively. FLT3/ITD+ cases showed a predominance of the bcr3 isoform (P = 0.008) and M3v morphology (P < 0.001). We found that all FLT3/ITD+ cases expressed CD2 (9 of 9) more frequently than that of FLT3/ITD− (1 of 16) (P < 0.001), while only one of the CD2+ cases (1 of 10, 10%) did not harbor FLT3/ITD, and all CD2+CD34+ cases (5 of 5, 100%) harbored FLT3/ITD. In addition, quantitative polymerase chain reaction analysis showed that FLT3 mRNA was more abundantly expressed in FLT3/ITD+ than that in FLT3/ITD− (P = 0.025), while there was no difference between D835(I836)+ and D835(I836)− with regards to aberrant surface-antigen expression, expression levels of FLT3 mRNA, M3v morphology, and the bcr3 isoform of PML-RARa mRNA. This study demonstrates that the presence of FLT3/ITD, but not D835 (I836), is closely related to aberrant CD2 expression and high expression levels of FLT3 mRNA. Our findings also suggest that FLT3/ITD as a secondary genetic event may block differentiation at the immature stage of APL.

A quantitative nested reverse transcriptase polymerase chain reaction (QN‐RT‐PCR) method was developed using a plasmid cDNA containing the AML1/ETO (MTG8) fusion transcript from Kasumi‐1 cells, an acute‐myelogenous leukemia cell line with the t(8;21) translocation. In this method, the plasmid was detectable at a concentration of 10⁻¹⁷ m. The fusion transcript in a mixture of 10⁷ Rice94 (Burkitt lymphoma cell line) cells containing two Kasumi‐1 cells was detectable at 10⁻¹⁷ m. In a previously published real‐time PCR method, the plasmid containing the fusion transcript was detectable at 10⁻¹⁶ m or higher, and 20 or more Kasumi‐1 cells were detectable in 10⁷ Rice94 cells. Thus, this QN‐RT‐PCR method is more sensitive than the real‐time PCR. When the same samples were examined by real‐time PCR and our QN‐RT‐PCR method, in one patient in clinical remission after chemotherapy and allogeneic‐bone marrow transplantation (BMT), the transcript was detected by QN‐RT‐PCR 60 days prior to hematological relapse, in contrast to 10 days before hematological relapse by real‐time PCR. The transcript level was below 10⁻¹⁷ m (undetectable) with this QN‐RT‐PCR in patients in clinical remission after chemotherapy and BMT, while it was 10⁻¹⁵–10⁻¹⁶ m in patients in clinical remission after chemotherapy alone. The quantitative difference of the transcript level in minimal residual disease (MRD) between these two different types of clinical remission was estimated to be at least 10²‐fold. This QN‐RT‐PCR method is useful for predicting hematological relapse and for quantitatively estimating MRD in different types of clinical remission.

Accumulation of advanced glycation end products (AGEs) of the Maillard reaction has been implicated in the pathogenesis of diabetes and its complications. Connarus ruber has been used as a folk remedy for several diseases, including diabetes; however, its underlying mechanism has not yet been investigated. This study investigated the effects of C. ruber extract against glycation on collagen-linked AGEs in vitro and streptozotocin-induced diabetic rats (STZ-DM rats) in vivo. The antiglycation activities of C. ruber extract and aminoguanidine (AG) were examined using a collagen glycation assay kit. Nonfluorescent AGE, N^ε-carboxymethyl lysine (CML), N^ω-carboxymethyl arginine, and N^ε-carboxyethyl lysine levels were measured via electrospray ionization-liquid chromatography-tandem mass spectrometry. The effect of the extract on the cytotoxicity of methylglyoxal (MG), a precursor of AGEs, was examined in HL60 cells. STZ-DM rats were treated with the extract for 4 wk, and the effect was assessed using biochemical markers in the serum and CML-positive cells in renal tissues. C. ruber extract dose-dependently inhibited the glycation of collagen and formation of nonfluorescent AGEs, which was comparable to AG, and it significantly attenuated MG-induced cytotoxicity in HL60 cells. Furthermore, the glycated albumin levels in STZ-DM rats decreased, the increase in serum lipid levels was reversed, and immunohistochemistry demonstrated that CML deposition in the glomerulus of STZ-DM rats significantly decreased. Although further studies are needed, C. ruber could be a potential therapeutic for preventing and progressing many pathological conditions, including diabetes. Received: 09 June 2021 Accepted after revision: 08 November 2021 Publication Date: 03 December 2021 (online) © 2021. Thieme. All rights reserved. Georg Thieme Verlag KG Rüdigerstraße 14, 70469 Stuttgart, Germany

Background: Glycation, a non-enzymatic reaction between sugars and biomolecules, leads to the formation of advanced glycation end-products (AGEs), which are implicated in the progression of chronic diseases. Connarus ruber (Poepp.) Planch (C. ruber), a traditional medicinal plant used for diabetes, has shown anti-glycation activity. This study aimed to identify the active components in C. ruber extract and elucidate their anti-glycation mechanisms. Methods: Using NMR and LC-MS analyses, we identified epicatechin and procyanidin A2 as major polyphenolic constituents. Collagen glycation assays were performed to evaluate the inhibitory effects of these compounds on fructose- and glyceraldehyde (GA)-induced glycation. Additionally, their cytoprotective effects were assessed using GA-induced cytotoxicity assays in dental pulp stem cells (DPSCs). Results: Both epicatechin and procyanidin A2 inhibited fructose- and GA-induced glycation in a dose-dependent manner, showing greater efficacy than aminoguanidine. Furthermore, these compounds significantly alleviated GA-induced cytotoxicity in DPSCs. Conclusions: These findings suggest that epicatechin and procyanidin A2 are candidate contributors to the anti-glycation and cytoprotective effects of C. ruber. The results support the potential of C. ruber extract as a source of therapeutic agents for glycation-related diseases and for enhancing stem cell viability.