1. Viruses and Encephalitis
PHM142
September 24, 2019
Amy (Yoo Im) Choi
Candice (Zinan) Lin
Aiwen Qian
PHM Fall 2019
Instructor: Chesa Dojo Soeandy
Coordinator: Jeffrey Henderson

2. What is encephalitis? Inflammation of cranial tissue 2 major subtypes:
Infection-related
Autoimmune-mediated
Symptoms:
Ex. focal neurological deficits, seizures
Hard to identify etiological agent
Encephalitis means inflammation of the cranial tissues and it is a rare but a very serious neurologic condition. The etiology of encephalitis can be broadly divided into 2 major subtypes: the first one is infection-related encephalitis, which is directly caused by pathogenic viral, bacterial, or parasitic agents; the second one is autoimmune-mediated encephalitis, which is mediated by an abnormal immune response. This can be triggered by a recent viral infection or vaccination. We will be focusing on the infection-related acute encephalitis, which is most commonly associated with viruses. After the virus invades the CNS, a focal inflammatory and necrotizing lesion is observed in the temporal lobes (as shown in this MRI of the brain); the inflammation can also be in the orbital frontal cortex, and limbic structures. Severe symptoms of acute encephalitis include, altered level of consciousness, focal neurological deficits (ex. Speech impairment and behavioural disturbance) and possibly seizure. In addition, diagnosing viral encephalitis can be challenging since almost 70% cases are left without an etiologic agent identified.

3. Viral replication
DNA viruses: Herpes Simplex Virus (HSV) 1, HSV2, adenovirus
RNA viruses: influenza, enterovirus, measles, rubella, mumps, rabies, arboviruses
Haematogenous route - Ex. arboviruses - affects blood-brain barrier
Arbovirus ex. West nile encephalitis virus
Virus first attaches itself to receptors on the cell membrane of the host cell using the proteins on the surface of its envelope. The contents of the virus are then released into the cytosol via endocytosis. Viral reverse transcriptase then copies RNA to form double stranded DNA. Then, the viral DNA is integrated into the host cell’s DNA, resulting in the production of viral proteins and structures. The new viral DNA and enzymes leave the cell via exocytosis, using the cell membrane to create the viral envelope. Now, it’s ready to infect other host cells.

4. Herpes Simplex Virus (HSV)
2 types: HSV-1 and HSV-2
Oral vs. sexually transmitted
Route of infection: HSV-1
Trigeminal nerve/Olfactory tract
Recurrent infection
Reactivation
Route of infection: HSV-2
Sexually transmitted
Vertically transmitted
Latent infections in sensory neurons
HSV-1 is the most common etiologic agent of acute encephalitis
HSV-1 can gain access in CNS through one of the following routes
o 1. Invasion via trigeminal nerve or olfactory tract after primary infection in the oropharynges
o 2. CNS penetration after recurrent viral infection (viral reactivation followed by viral spread)
o 3. CNS invasion after viral reactivation of a latent HSV in the brain
-> After CNS invasion, a focal inflammatory and necrotizing lesion is observed specially in the temporal lobes, orbital frontal cortex, and limbic structures
HSV-1 is normally associated with orofacial infections and HSV-2 usually causes genital infections and can be transmitted from infected mothers to neonates
o Both viruses establish latent infections in sensory neurons and, upon reactivation, cause lesions at or near point of entry into the body
HSV-2 is usually sexually transmitted and most genital HSV infections are caused by HSV-2
Focus on HSV
½ cases < 20 years old vs. ½ cases > 50 years old (b/c primary vs. latent infection)
HSV2 has latency in sacral dorsal route ganglia
-infection of CNS → subacute encephalitis, acute HSE. Pathway unknown
Crucial early diagnosis - clinical symptoms include inflammation and swelling of brain tissue
There are 2 types of Herpes Simplex virus - type 1 and 2. HSV-1 is by far the most common etiologic agent of acute encephalitis in the world. Almost 90% of all adult cases of HSV encephalitis are caused by HSV-1. As you can see on this image, HSV-1 is usually transmitted orally and associated with orofacial infection. It can gain access in the CNS in one of three ways: one route is by invasion via trigeminal nerve or olfactory tract after primary infection in the oropharynx; another route is by CNS penetration after recurrent viral infection, that is viral reactivation followed by viral spread; and finally, the third possible route is by viral reactivation of a latent HSV in the brain. In comparison, HSV-2 usually causes genital infections through sexual transmission. It can also be transmitted from infected mothers to neonates. However, both viruses can cause latent infections in sensory neurons and, upon reactivation, cause lesions at or near point of entry into the body.

5. Herpes simplex encephalitis (HSE)
Unknown pathway to CNS target
Early diagnosis crucial
Cerebrospinal fluid PCR:
98% sensitivity; 94% specificity
Faster vs. serologic testing
Monitoring therapy
Axoplasmic transport (HSV1):
Reactivation from trigeminal sensory ganglion → retrograde virus transport → oral herpes
HSV2 latency in dorsal route ganglia:
Infects CNS → subacute encephalitis, acute HSE
But diagnosis challenging b/c non-specific clinical symptoms
Initial clinical symptoms:
altered consciousness (can vary from drowsiness→ coma) and signs of diffuse cerebral dysfunction
diffuse and focal cerebral pathology, also meningitis (fever,headache)
PCR:
Other methods not widely used includes nucleic acid sequence-based amplification, branched-DNA assay
Retains sensitivity even after taking antiviral drugs
Detectable in CSF min. 1 week after symptoms commence
Higher copy number detected associated with more severe symptoms
Also for monitoring therapy effectiveness

6. Blood Brain Barrier (BBB) Disruption
HSV Infection
Intracellular Adhesion Molecule 1 (ICAM-1)
Matrix Metalloproteinases (MMPs)
Aquaporin 4 (AQP4)
Endothelial Nitric Oxide Synthase (eNOS)
Water into astrocyte
Water into basal membrane
Astrocyte apoptosis
Cleaves tight junction proteins
Nitric Oxide (NO)
Brain edema
com.myaccess.library.utoronto.ca/content/pdf/ %2Fs pdf
Although the pathway of HSE is unknown, there has been an association found between HSE and the disruptions to BBB, which gives an insight into the mechanism.
-ICAM-1 determines the endothelial barrier and it regulates leukocyte movement across endothelial cells. Therefore, an increase in ICAM-1 expression caused by HSV infection leads to an increase in leukocyte interaction with endothelial cells.
-HSV causes upregulation of MMP expression. MMPs are endopeptidases that cleave TJ proteins
-eNOS is upregulated. NO is a vasodilator which can increase endothelial cell permeability.
-HSV promotes the upregulation of AQP4 in the chronic phase (after 6 months)
-Astrocytes protect the brain against pathogens by secreting immune factors
Blood Brain Barrier (BBB) Disruption

7. Treatment: Acyclovir Mechanism of Action
Here we start with a host cell that has been infected by the virus. The blue molecules on the left represent the drug, acyclovir. Once it enters the cell, viral thymidine kinase will phosphorylate it. Then, GMP kinase phosphorylates it to form DP. The third phosphorylation is done by nucleoside diphosphate kinase. The later two kinases are cellular, whereas the first one was from the virus itself. Now the ACV-TP enters the nucleus. That’s the viral DNA polymerase, replicating DNA. The drug is then incorporated into the growing DNA, where it causes chain termination. So now, with a constant high concentration of the drug, the virus will stop replicating. And that treats the viral infection.

8. Treatment: Acyclovir Mechanism of Action
Dose - 10 mg/kg every 8 hours, days
Condense!!!
On the left, we have guanosine, which is made of guanine + deoxyribose. On the right, we have acyclovir, the guanosine analogue. The important thing to note here is the missing free 3’ hydroxyl group, which is needed for DNA synthesis. As seen previously in the video, ACV undergoes several phosphorylation steps. The phosphates are added to the 5’ hydroxyl end. Once it reaches the nucleus, it gets added to the growing DNA chain, and successfully terminates the chain because of the missing 3’ OH. Additionally, ACV-TP can deactivate the viral DNA polymerase. The specificity of the drug is what makes it very effective. One, ACV does not get phosphorylated by uninfected cells b/c of its high affinity to VTK. Second, ACV-TP is a more potent inhibitor of viral DNA polymerase than normal cells.

9. Summary Encephalitis Encephalitis = inflammation of cranial tissue
DNA or RNA viral causes
Most common viral cause of encephalitis: Herpes simplex virus
Herpes simplex virus (2 types)
HSV-1: oral-oral infection
HSV-2: sexually transmitted infection and vertical infection
Herpes simplex encephalitis
Unknown complete mechanism
CSF-PCR for diagnosis
Blood-brain barrier disruption associated with HSE
First line therapy for HSE: acyclovir (guanosine analogue)

10. References
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