Expanding structural diversity in a library of disulfide macrocycles through in-situ imide hydrolysis

We describe here an unorthodox approach to dynamic covalent chemistry in which the initially-unexpected in-situ hydrolysis of a bis-imide is employed to control the composition of a library of structurally diverse macrocycles. A single building block is used to generate a library of numerous disulfide-based architectures in a one-pot single-step process. The dual-stimuli method is based on simultaneous changes in pH and DMSO concentration to expand the structural diversity of the macrocyclic products. Mechanistic details of this complex process are investigated by the kinetics analysis. We delivered a facile strategy for the synthesis of water-soluble, multicomponent and dynamic macrocycles equipped with number of different functional groups, thus giving a prospect of their application in guest-driven phase transfer.

DMSO Concentrations up to 1% are Safe to be Used in the Zebrafish Embryo Developmental Toxicity Assay

Dimethyl sulfoxide (DMSO) is a popular solvent for developmental toxicity testing of chemicals and pharmaceuticals in zebrafish embryos. In general, it is recommended to keep the final DMSO concentration as low as possible for zebrafish embryos, preferably not exceeding 100 μL/L (0.01%). However, higher concentrations of DMSO are often required to dissolve compounds in an aqueous medium. The aim of this study was to determine the highest concentration of DMSO that can be safely used in our standardized Zebrafish Embryo Developmental Toxicity Assay (ZEDTA). In the first part of this study, zebrafish embryos were exposed to different concentrations (0–2%) of DMSO. No increase in lethality or malformations was observed when using DMSO concentrations up to 1%. In a follow-up experiment, we assessed whether compounds that cause no developmental toxicity in the ZEDTA remain negative when dissolved in 1% DMSO, as false positive results due to physiological disturbances by DMSO should be avoided. To this end, zebrafish embryos were exposed to ascorbic acid and hydrochlorothiazide dissolved in 1% DMSO. Negative control groups were also included. No significant increase in malformations or lethality was observed in any of the groups. In conclusion, DMSO concentrations up to 1% can be safely used to dissolve compounds in the ZEDTA.

Evaluating the affect of dimethyl sulfoxide on the viablity of H9C2 cardiomyocytes

Objective: The study was conducted to evaluate the affect of Dimethyl Sulfoxide (DMSO) concentration on the viability of H9C2 cells under the different cultural conditions. Methods: H9C2 cells were cultured under normal conditions and subjected to hypoxia/reoxygenation model. DMSOat the doses of 0,001÷10% (v/v) was added to the cultural medium during normal culture period and reoxygenation period. Cellular viability of the experimental groups was assessed by using CCK-8 kit. Results: The results indicated that the viability of H9C2 cardiomyocytes was stable in the different culture media supplied with DMSO at the doses of0,001÷0,5% (v/v). Meanwhile, supplementation of DMSO at the doses of1% and 2% significantly decreased the survival rate of H9C2 cells (v/v,p<0,05). Conclusion: The affect of DMSO on H9C2 cardiomyocytes is dose-dependant maner.

Freezing Medium Containing 5% DMSO Enhances the Cell Viability and Recovery Rate After Cryopreservation of Regulatory T Cell Products ex vivo and in vivo

Cell therapies have significant therapeutic potential in diverse fields including regenerative medicine, transplantation tolerance, and autoimmunity. Within these fields, regulatory T cells (Treg) have been deployed to ameliorate aberrant immune responses with great success. However, translation of the cryopreservation strategies employed for other cell therapy products, such as effector T cell therapies, to Treg therapies has been challenging. The lack of an optimized cryopreservation strategy for Treg products presents a substantial obstacle to their broader application, particularly as administration of fresh cells limits the window available for sterility and functional assessment. In this study, we aimed to develop an optimized cryopreservation strategy for our CD4+CD25+Foxp3+ Treg clinical product. We investigate the effect of synthetic or organic cryoprotectants including different concentrations of DMSO on Treg recovery, viability, phenotype, cytokine production, suppressive capacity, and in vivo survival following GMP-compliant manufacture. We additionally assess the effect of adding the extracellular cryoprotectant polyethylene glycol (PEG), or priming cellular expression of heat shock proteins as strategies to improve viability. We find that cryopreservation in serum-free freezing medium supplemented with 10% human serum albumin and 5% DMSO facilitates improved Treg recovery and functionality and supports a reduced DMSO concentration in Treg cryopreservation protocols. This strategy may be easily incorporated into clinical manufacture protocols for future studies.

Expanding Structural Diversity In A Library Of Disulfide Macrocycles Through In-Situ Imide Hydrolysis

We describe here an unorthodox approach to dynamic covalent chemistry in which the initially-unexpected in-situ hydrolysis of a bis-imide component is used to control the composition of a library of structurally diverse macrocycles. The dual-stimuli method is based on simultaneous changes in pH and DMSO concentration to expand the diversity of the macrocyclic architectures.

THE EFFECT OF DIMETHYLSULFOXIDE ON THE FLUORESCENCE PROPERTIES OF SOME 4-HYDROXYQUINOLINES

Fluorescence properties of 4-hydroxy-2-methylquinoline (1) and 2-(5-mercapto-1,3,4-oxadiazol-2-yl)-6-methylquinoline-4-ol (2) were studied in dimethylsulfoxide (DMSO) aqueous solutions. The fluorescence properties of 1 and 2 exhibit substantial dependence on the DMSO concentration. The fluorescence quantum yield $(\Phi_f)$ of 1 decreases upon adding DMSO due to the shift in the keto-enol (E) tautomeric equilibrium toward E form․ On the contrary 2 demonstrates a tendency of increase of $\Phi_f$ upon adding DMSO due to intermolecular charge transfer from DMSO to the aromatic ring of quinoline, which increases the electron density on the ring and hence the fluorescence efficiency.

Numerical Study of Heat and Mass Transfer during Cryopreservation Process with Application of Directed Interval Arithmetic

In the present paper, numerical modelling of heat and mass transfer proceeding in a two-dimensional axially symmetrical articular cartilage sample subjected to a cryopreservation process is presented. In the model under consideration, interval parameters were assumed. The heat transfer process is described using the Fourier interval equation, while the cryoprotectant transport (DMSO) across the cell membrane is analyzed using a two-parameter model taking into account the simulation of the water volume in the chondrocytes and the change in DMSO concentration over time. The liquidus tracking (LT) protocol introduced by Pegg et al. was used to model the cryopreservation process. This procedure divides the heating and cooling phases into eight and seven steps, respectively, allowing precise regulation of temperature and cryoprotectant (CPA) concentration of bathing solutions. This protocol protects chondrocytes from ice crystal, osmotic stress, and electrolyte damage. The obtained interval concentrations of cryoprotectant in chondrocytes were compared with previous simulations obtained using the deterministic model and they are mostly in agreement with the simulation data.

Post-Cryo Survival of Rat Testicular Interstitial Cells in Hydroxyethyl Starch-Based Media

The development of cryoprotective serum-free or xeno-free media is required for safe use of cryopreserved testicular material for transplantation. In this study, the solutions containing 10% fetal bovine serum (FBS) or 5 mg/ml bovine serum albumin (BSA) did not significantly enhance the general survival of interstitial cells (ICs) after cryopreservation but increase their metabolic activity and steroid producing cell (HSD+-cells) survival. The use of 50 and 100 mg/ml hydroxyethyl starch (HES) in DMSO-based cryoprotective solutions instead of BSA or FBS enabled the improvement of the IC general survival and the survival of HSD+-cells. The use of HES supplemented media allowed to decrease the dimethyl sulfoxide (DMSO) concentration from 1.4 to 0.7 M and to preserve the amount and metabolic activity of ICs. Thus, designing cryoprotective media containing DMSO and HES can facilitate the formulation of serum-free solutions for cryopreservation that in turn paves a way for implementation of the use of cryopreserved material for practical medicine.

Non-Uniform Distribution of Cryoprotecting Agents in Rice Culture Cells Measured by CARS Microscopy

Cryoprotectants allow cells to be frozen in liquid nitrogen and cryopreserved for years by minimizing the damage that occurs in cooling and warming processes. Unfortunately, how the specific cryoprotectants keep the cells viable through the cryopreservation process is not entirely evident. This contributes to the arduous process of optimizing cryoprotectant formulations for each new cell line or species that is conserved. Coherent anti-Stokes Raman scattering microscopy facilitates the visualization of deuterated cryoprotectants within living cells. Using this technique, we directly imaged the location of fully deuterated dimethyl sulfoxide (d6-DMSO), the deuterated form of a commonly used cryoprotectant, DMSO, within rice suspension cells. This work showed that d6-DMSO does not uniformly distribute throughout the cells, rather it enters the cell and sequesters within organelles, changing our understanding of how DMSO concentration varies within the cellular compartments. Variations in cryoprotectant concentration within different cells and tissues will likely lead to differing protection from liquid nitrogen exposure. Expanding this work to include different cryoprotectants and mixtures of cryoprotectants is vital to create a robust understanding of how the distributions of these molecules change when different cryoprotectants are used.