Abstract: Duchenne muscular dystrophy (DMD) is a severe, progressive muscle disease caused by mutations in the dystrophin gene. The majority of DMD mutations are deletions that prematurely terminate the dystrophin protein. Deletions of exon 50 of the dystrophin gene are among the most common single exon deletions causing DMD. Such mutations can be corrected by skipping exon 51, thereby restoring the dystrophin reading frame. Using clustered regularly interspaced short palindromic repeats/CRISPR-associated 9 (CRISPR/Cas9), we generated a DMD mouse model by deleting exon 50. These ΔEx50 mice displayed severe muscle dysfunction, which was corrected by systemic delivery of adeno-associated virus encoding CRISPR/Cas9 genome editing components. We optimized the method for dystrophin reading frame correction using a single guide RNA that created reframing mutations and allowed skipping of exon 51. In conjunction with muscle-specific expression of Cas9, this approach restored up to 90% of dystrophin protein expression throughout skeletal muscles and the heart of ΔEx50 mice. This method of permanently bypassing DMD mutations using a single cut in genomic DNA represents a step toward clinical correction of DMD mutations and potentially those of other neuromuscular disorders.

Abstract: The development of CRISPR-Cas9 RNA-guided genome editing has transformed biomedical research. Most applications reported thus far rely upon the Cas9 protein from Streptococcus pyogenes SF370 (SpyCas9). With many RNA guides, SpyCas9 can induce significant levels of unintended mutations at near-cognate sites, necessitating substantial efforts toward the development of strategies to minimize off-target activity. Although the genome-editing potential of thousands of other Cas9 orthologs remains largely untapped, it

Abstract: INTRODUCTION: Tacrolimus (Tac) is the cornerstone of immunosuppressive therapy after solid organ transplantation and will probably remain so. Excluding belatacept, no new immunosuppressive drugs were registered for the prevention of acute rejection during the last decade. For several immunosuppressive drugs, clinical development halted because they weren’t sufficiently effective or more toxic. Areas covered: Current methods of monitoring Tac treatment, focusing on traditional therapeutic drug monitoring (TDM), controversies surrounding TDM, novel matrices, pharmacogenetic and pharmacodynamic monitoring are discussed. Expert opinion: Due to a narrow therapeutic index and large interpatient pharmacokinetic variability, TDM has been implemented for individualization of Tac dose to maintain drug efficacy and minimize the consequences of overexposure. The relationship between predose concentrations and the occurrence of rejection or toxicity is controversial. Acute cellular rejection also occurs when the Tac concentration is within the target range, suggesting that Tac whole blood concentrations don’t necessarily correlate with pharmacological effect. Intracellular Tac, the unbound fraction of Tac or pharmacodynamic monitoring could be better biomarkers/tools for adequate Tac exposure – research into this has been promising. Traditional TDM, perhaps following pre-emptive genotyping for Tac-metabolizing enzymes, must suffice for a few years before these strategies can be implemented in clinical practice.

Abstract: Hemolytic uremic syndrome (HUS) is an unrare and severe thrombotic microangiopathy (TMA) caused by several pathogenetic mechanisms among which Shiga toxin-producing Escherichia coli infections and complement dysregulation are the most common. However, very rarely and particularly in neonates and infants, disorders of cobalamin metabolism (CblC) can present with or be complicated by TMA. Herein we describe a case of atypical HUS (aHUS) related to CblC disease which first presented in a previously healthy boy at age of 13.6 years. The clinical picture was initially dominated by nephrotic range proteinuria and severe hypertension followed by renal failure. The specific treatment with high dose of hydroxycobalamin rapidly obtained the remission of TMA and the complete recovery of renal function. We conclude that plasma homocysteine and methionine determinations together with urine organic acid analysis should be included in the diagnostic work-up of any patient with TMA and/or nephrotic syndrome regardless of age.

Abstract: Magnesium gluconate is a potent antioxidant used for the prevention and treatment of hypomagnesia. The current research was aimed to investigate the impact of The Trivedi Effect® – Energy of Consciousness Healing Treatment (Biofield Energy Healing Treatment) on magnesium gluconate for the change in the physicochemical, structural, thermal and behavioral properties using PXRD, PSD, FT-IR, UV-vis spectroscopy, TGA, and DSC analysis. Magnesium gluconate was divided into two parts – one part was control, while another part was treated with The Trivedi Effect® – Energy of Consciousness Healing Treatment (Biofield Energy Healing Treatment) remotely by twenty renowned Biofield Energy Healers and defined as The Trivedi Effect® treated sample. The PXRD analysis exhibited that the crystallite size of the treated sample was remarkably changed from range -37.52% to 180.14% compared with the control sample. The average crystallite size was reduced in the treated sample by 0.11% compared with the control sample. PSD analysis revealed that the particle sizes in the treated sample at d10 and d50 values were significantly decreased by 11.36% and 8.25%, respectively, while the particle size at d90 was increased in the treated sample by 2.43% compared with the control sample. The treated sample’s surface area was significantly increased by 9.54% compared with the control sample. The FT-IR and UV-vis analysis showed that the structure of the magnesium gluconate remained the same in both the treated and control samples. The TGA analysis revealed the four steps thermal degradation of the both samples and the total weight loss of the treated sample was reduced by 2.32% compared with the control sample. This result indicated that the thermal stability of the treated sample was improved compared with the control sample. The DSC analysis revealed that the melting temperature of the treated sample (171.05°C) was decreased by 0.17% compared with the control sample (171.34°C). The latent heat of fusion was decreased by 1.16% in the treated sample compared with the control sample. The current study infers that The Trivedi Effect® – Energy of Consciousness Healing Treatment (Biofield Energy Healing Treatment) might lead to a new polymorphic form of magnesium gluconate, which would be more soluble, bioavailable, and thermally stable compared with the untreated compound. Hence, The Trivedi Effect® Treated magnesium gluconate would be very useful to design better nutraceutical and/or pharmaceutical formulations that might offer better therapeutic responses against inflammatory diseases, immunological disorders, stress, aging and other chronic infections.