The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRISPR-associated sequence (CRISPR/Cas) system is a cutting-edge genome-editing tool employed to explore the functions of normal and disease-related genes. The CRISPR/Cas system has a remarkable diversity in the composition and architecture of genomic loci and Cas protein sequences. Owing to its excellent efficiency and specificity, this system adds an outstanding dimension to biomedical research on genetic manipulation of eukaryotic cells. However, safe, efficient, and specific delivery of this system to target cells and tissues and their off-target effects are considered critical bottlenecks for the therapeutic applications. Recently discovered anti-CRISPR proteins (Acr) play a significant role in limiting the effects of this system. Acrs are relatively small proteins that are highly specific to CRISPR variants and exhibit remarkable structural diversity. The in silico approaches, crystallography, and cryo-electron microscopy play significant roles in elucidating the mechanisms of action of Acrs. Acrs block the CRISPR/Cas system mainly by employing four mechanisms: CRISPR/Cas complex assembly interruption, target-binding interference, target cleavage prevention, and degradation of cyclic oligonucleotide signaling molecules. Engineered CRISPR/Cas systems are frequently used in gene therapy, diagnostics, and functional genomics. Understanding the molecular mechanisms underlying Acr action may help in the safe and effective use of CRISPR/Cas tools for genetic modification, particularly in the context of medicine. Thus, attempts to regulate prokaryotic CRISPR/Cas surveillance complexes will advance the development of antimicrobial drugs and treatment of human diseases. In this review, recent updates on CRISPR/Cas systems, especially CRISPR/Cas9 and Acrs, and their novel mechanistic insights are elaborated. In addition, the role of Acrs in the novel applications of CRISPP/Cas biotechnology for precise genome editing and other applications is discussed.

The genome editing approach by clustered regularly interspaced short palindromic repeats (CRISPR)/associated protein 9 (CRISPR/Cas9) is a revolutionary advancement in genetic engineering. Owing to its simple design and powerful genome-editing capability, it offers a promising strategy for the treatment of different infectious, metabolic, and genetic diseases. The crystal structure of Streptococcus pyogenes Cas9 (SpCas9) in complex with sgRNA and its target DNA at 2.5 Å resolution reveals a groove accommodating sgRNA:DNA heteroduplex within a bilobate architecture with target recognition (REC) and nuclease (NUC) domains. The presence of a PAM is significantly required for target recognition, R-loop formation, and strand scission. Recently, the spatiotemporal control of CRISPR/Cas9 genome editing has been considerably improved by genetic, chemical, and physical regulatory strategies. The use of genetic modifiers anti-CRISPR proteins, cell-specific promoters, and histone acetyl transferases has uplifted the application of CRISPR/Cas9 as a future-generation genome editing tool. In addition, interventions by chemical control, small-molecule activators, oligonucleotide conjugates and bioresponsive delivery carriers have improved its application in other areas of biological fields. Furthermore, the intermediation of physical control by using heat-, light-, magnetism-, and ultrasound-responsive elements attached to this molecular tool has revolutionized genome editing further. These strategies significantly reduce CRISPR/Cas9’s undesirable off-target effects. However, other undesirable effects still offer some challenges for comprehensive clinical translation using this genome-editing approach. In this review, we summarize recent advances in CRISPR/Cas9 structure, mechanistic action, and the role of small-molecule activators, inhibitors, promoters, and physical approaches. Finally, off-target measurement approaches, challenges, future prospects, and clinical applications are discussed.

Background: Diabetes mellitus (DM) is a major health burden affecting 537 million adults worldwide, characterized by chronic metabolic disorder and various complications. This case control study aimed to assess the impact of type 2 diabetes mellitus (T2DM), including hyperglycemia levels, on hematological parameters and complete blood count (CBC) derived parameters. Methods: A total of 250 known diabetic patients from the Jazan Diabetic Center, Saudi Arabia, between January 2021 and December 2022, along with 175 healthy adult controls were recruited from Jazan Hospital’s blood donation center. Demographic characteristics, medical histories, and relevant factors such as gender, age, BMI, treatment, disease duration, and comorbidities were collected with informed consent. Results: The results of the red blood cell (RBC) count, RBC indices, and mean platelet volume showed significant differences between patients and controls, while the white cell (WBC) and platelet count were comparable between the two groups. CBC-derived parameters, especially neutrophil/lymphocyte ratio (NLR), and platelet/neutrophil ratio (PNR) exhibited significant differences. Conclusion: CBC and derived parameters serve as inexpensive tools for T2DM patients monitoring, indicating early blood cell alterations and potential development of anemia. Further studies are needed to explore their role in predicting T2DM pathogenesis and progression, aiming to reduce severe complications, mortality and morbidity.

Purpose: Sickle cell disease (SCD) and thalassemia are common inherited blood disorders in Saudi Arabia, especially in Jazan Province. Patients with these disorders require multiple blood transfusions, which may lead to alloimmunization because of mismatched blood group antigens. In this study, we examined the alloimmunization and autoimmunization rates in patients with SCD and thalassemia together with the involved antibodies. Patients and Methods: A cross-sectional study was conducted to review the transfusion history records of patients with SCD and thalassemia at Prince Mohammed bin Nasser Hospital, Jazan Province, Saudi Arabia. Results: Four-hundred thirty-eight patients (385 with SCD, 52 with β-thalassemia, and 1 with α-thalassemia) were received leukoreduced red cell transfusions. The alloimmunization and autoimmunization rates in patients with SCD were 12.98% and 0.52%, respectively. In patients with thalassemia, the alloimmunization and autoimmunization rates were 13.21% and 3.77%, respectively. The most prevalent antibodies in the study population were anti-E (17.19%) and anti-K (14.06%). Conclusion: The alloimmunization and autoimmunization rates were determined in patients with SCD and thalassemia in Jazan Province, Saudi Arabia. The results highlight the need for extended phenotyping to include ABO, RH (D, C, c, E, e), K, Fy^a, Fy^b, Jk^a and Jk^b antigens in the screening panel. This will benefit patients to ensure better transfusion practices.

Advanced mitochondrial multi-omics indicate a multi-facet involvement of mitochondria in the physiology of the cell, changing the perception of mitochondria from being just the energy-generating organelles to organelles that highly influence cell structure, function, signaling, and cell fate. This sets mitochondrial dysfunction in the centerstage of numerous acquired and genetic diseases. Sickle cell disease is also being increasingly associated with mitochondrial anomalies and the pathophysiology of sickle cell disease finds mitochondria at crucial intersections in the pathological cascade. Altered mitophagy, increased ROS, and mitochondrial DNA all contribute to the condition and its severity. Such mitochondrial aberrations lead to consequent mitochondrial retention in red blood cells in sickle cell diseases, increased oxidation in the cellular environment, inflammation, worsened vaso-occlusive crisis, etc. There are increasing studies indicating mitochondrial significance in sickle cell disease, consequently providing an opportunity to target it for improving the outcomes of treatment. Identification of the impaired mitochondrial attributes in sickle cell disease and their modulation by therapeutic interventions can impart a better management of the disease. This review aims to describe the mitochondria in the perspective of sicke cell disease so as to provide the reader an overview of the emerging mitochondrial stance in sickle cell disease.

Purpose: Knowledge of the prevalence of blood group antigens in a given population is important for the prevention of hemolytic reactions. The MNS blood group system (002) has four polymorphic antigens—M, N, S, and s. Anti-S and anti-s antibodies may result in immediate and delayed hemolytic transfusion reactions, and hemolytic disease of the fetus and newborn may occur. The present study investigated the frequencies of the main antigens and phenotypes of the MNS blood group system. Subjects and Methods: We randomly obtained 149 samples from anonymous Saudi blood donors living in Jazan Province. Serotyping was conducted using a gel card to investigate (M, N, S, and s) antigens and phenotypes. Results: The frequencies of MNS antigens were as follows: M = 89.26%, N = 51.67%, S = 61.07%, and s = 82.55%. Regarding the MNS phenotypes, nine phenotypes were observed in the study population. The most common phenotype was M+N–S+s+ (n = 36, 24.16%), in contrast to the least common phenotype M+N–S–s– (n = 1, 0.67%). The prevalence of the MNS phenotypes in the current study population was highly and significantly different from that in Europeans (P = 0.044) and African Americans (P = 0.000). Conclusion: In summary, this study reports the frequencies of the MNS antigens and phenotypes in Jazan Province, Saudi Arabia. The most common phenotype was M+N–S+s+, whereas the least observed phenotype was M+N–S–s–. The outcomes of this study may assist the blood banks in Jazan Province to establish an extended phenotyping protocol including the MNS antigens, in particular S and s antigens, to preclude any alloimmunization events.