ABSTRACT
The CRISPR/Cas9 gene editing method is comprised of the guide RNA (gRNA) to target a specific DNA sequence for cleavage and the Cas9 endonuclease for introducing breaks in the double-stranded DNA identified by the gRNA. Co-expression of both a multiplex of HIV-1-specific gRNAs and Cas9 in cells results in the modification and/or excision of the segment of viral DNA, leading to replication-defective virus. In this study, we have personalized the activity of CRISPR/Cas9 by placing the gene encoding Cas9 under the control of a minimal promoter of HIV-1 that is activated by the HIV-1 Tat protein. We demonstrate that functional activation of CRISPR/Cas9 by Tat during the course of viral infection excises the designated segment of the integrated viral DNA and consequently suppresses viral expression. This strategy was also used in a latently infected CD4+ T-cell model after treatment with a variety of HIV-1 stimulating agents including PMA and TSA. Controlled expression of Cas9 by Tat offers a new strategy for safe implementation of the Cas9 technology for ablation of HIV-1 at a very early stage of HIV-1 replication during the course of the acute phase of infection and the reactivation of silent proviral DNA in latently infected cells.
Subject(s)
Gene Editing , Gene Expression Regulation, Viral , HIV-1/genetics , 3' Untranslated Regions , 5' Untranslated Regions , Base Sequence , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/virology , Clustered Regularly Interspaced Short Palindromic Repeats , Genes, Viral , HIV Long Terminal Repeat , HIV-1/physiology , Humans , Jurkat Cells , Promoter Regions, Genetic , Sequence Homology, Nucleic Acid , Virus ReplicationABSTRACT
The study of neurological infections by viruses defines the field of neurovirology, which has emerged in the last 30 years and was founded upon the discovery of a number of viruses capable of infecting the human nervous system. Studies have focused on the molecular and biological basis of viral neurological diseases with the aim of revealing new therapeutic options. The first studies of neurovirological infections can be traced back to the discovery that some viruses have an affinity for the nervous system with research into rabies by Louis Pasteur and others in the 1880s. Today, the immense public health impact of neurovirological infections is illustrated by diseases such as neuroAIDS, progressive multifocal leukoencephalopathy, and viral encephalitis. Recent research has seen the development of powerful new techniques for gene editing that promise revolutionary opportunities for the development of novel therapeutic options. In particular, clustered regulatory interspaced short palindromic repeat-associated 9 system provides an effective, highly specific and versatile tool for targeting DNA viruses that are beginning to allow the development of such new approaches. In this short review, we discuss these recent developments, how they pertain to neurological infections, and future prospects.
Subject(s)
CRISPR-Cas Systems , Central Nervous System Viral Diseases/genetics , Central Nervous System Viral Diseases/therapy , Gene Editing/methods , Genetic Therapy/methods , Encephalitis, Herpes Simplex/genetics , Encephalitis, Herpes Simplex/therapy , HIV Infections/genetics , HIV Infections/therapy , Herpesvirus 1, Human/genetics , Humans , JC Virus/genetics , Leukoencephalopathy, Progressive Multifocal/genetics , Leukoencephalopathy, Progressive Multifocal/therapyABSTRACT
This case report analyses the anesthetic technique and early postoperative treatment of a five months old baby with congenital aortic stenosis. Administration of cardiotonics and diuretics was necessary before surgery because of severe heart failure. The patient was anesthetized with halothane in pure oxygen and ventilated by means of an Engström ventilator. Surgical treatment consisted of valvulotomy under extracorporeal circulation and normothermia. Recovery was uneventful.
