Lauren D. Martin

Propofol-induced apoptosis of neurones and oligodendrocytes in fetal and neonatal rhesus macaque brain

BACKGROUND Exposure of the fetal or neonatal non-human primate (NHP) brain to isoflurane or ketamine for 5 h causes widespread apoptotic degeneration of neurones, and exposure to isoflurane also causes apoptotic degeneration of oligodendrocytes (OLs). The present study explored the apoptogenic potential of propofol in the fetal and neonatal NHP brain. METHOD Fetal rhesus macaques at gestational age 120 days were exposed in utero, or postnatal day 6 rhesus neonates were exposed directly for 5 h to propofol anaesthesia (n=4 fetuses; and n=4 neonates) or to no anaesthesia (n=4 fetuses; n=5 neonates), and the brains were systematically evaluated 3 h later for evidence of apoptotic degeneration of neurones or glia. RESULTS Exposure of fetal or neonatal NHP brain to propofol caused a significant increase in apoptosis of neurones, and of OLs at a stage when OLs were just beginning to myelinate axons. Apoptotic degeneration affected similar brain regions but to a lesser extent than we previously described after isoflurane. The number of OLs affected by propofol was approximately equal to the number of neurones affected at both developmental ages. In the fetus, neuroapoptosis affected particularly subcortical and caudal regions, while in the neonate injury involved neocortical regions in a distinct laminar pattern and caudal brain regions were less affected. CONCLUSIONS Propofol anaesthesia for 5 h caused death of neurones and OLs in both the fetal and neonatal NHP brain. OLs become vulnerable to the apoptogenic action of propofol when they are beginning to achieve myelination competence.

Isoflurane exposure leads to apoptosis of neurons and oligodendrocytes in 20- and 40-day old rhesus macaques

Previously we reported that a 5-hour exposure of 6-day-old (P6) rhesus macaques to isoflurane triggers robust neuron and oligodendrocyte apoptosis. In an attempt to further describe the window of vulnerability to anesthetic neurotoxicity, we exposed P20 and P40 rhesus macaques to 5h of isoflurane anesthesia or no exposure (control animals). Brains were collected 3h later and examined immunohistochemically to analyze neuronal and glial apoptosis. Brains exposed to isoflurane displayed neuron and oligodendrocyte apoptosis distributed throughout cortex and white matter, respectively. When combining the two age groups (P20+P40), the animals exposed to isoflurane had 3.6 times as many apoptotic cells as the control animals. In the isoflurane group, approximately 66% of the apoptotic cells were oligodendrocytes and 34% were neurons. In comparison, in our previous studies on P6 rhesus macaques, approximately 52% of the dying cells were glia and 48% were neurons. In conclusion, the present data suggest that the window of vulnerability for neurons is beginning to close in the P20 and P40 rhesus macaques, but continuing for oligodendrocytes.

Lithium Protects Against Anaesthesia Neurotoxicity In The Infant Primate Brain.

Exposure of infant animals, including non-human primates (NHPs), to anaesthetic drugs causes apoptotic death of neurons and oligodendrocytes (oligos) and results in long-term neurodevelopmental impairment (NDI). Moreover, retrospective clinical studies document an association between anaesthesia exposure of human infants and significant increase in NDI. These findings pose a potentially serious dilemma because millions of human infants are exposed to anaesthetic drugs every year as part of routine medical care. Lithium (Li) at clinically established doses is neuroprotective in various cerebral injury models. We therefore investigated whether Li also protects against anaesthesia neurotoxicity in infant NHPs. On postnatal day 6 NHPs were anaesthetized with the widely used anaesthetic isoflurane (ISO) for 5 h employing the same standards as in a human pediatric surgery setting. Co-administration of Li completely prevented the acute ISO-induced neuroapoptosis and significantly reduced ISO-induced apoptosis of oligodendroglia. Our findings are highly encouraging as they suggest that a relatively simple pharmacological manipulation might protect the developing primate brain against the neurotoxic action of anaesthetic drugs while not interfering with the beneficial actions of these drugs. Further research is needed to determine Li’s potential to prevent long-term NDI resulting from ISO anaesthesia, and to establish its safety in human infants.

Isoflurane exposure for three hours triggers apoptotic cell death in neonatal macaque brain

Background Retrospective clinical studies suggest there is a risk for neurodevelopmental impairment following early childhood exposure to anaesthesia. In the developing animal brain, including those of non-human primates (NHPs), anaesthetics induce apoptotic cell death. We previously reported that a 5 h isoflurane (ISO) exposure in infant NHPs increases apoptosis 13-fold compared with control animals. However, the majority of paediatric surgeries requiring anaesthesia are of shorter durations. We examined whether 3 h ISO exposure similarly increases neuroapoptosis in the NHP developing brain. Methods Six-day-old NHP infants ( Macaca mulatta ) were exposed to 3 h of a surgical plane of ISO ( n =6) or to room air ( n =5). Following exposure, NHP brains were screened for neuronal and oligodendrocyte apoptosis using activated caspase-3 immunolabelling and unbiased stereology. Results ISO treatment increased apoptosis (neurones + oligodendrocyte) to greater than four times that in the control group [mean density of apoptotic profiles: 57 (SD 22) mm -3 vs 14 (SD 5.2) mm -3 , respectively]. Oligodendrocyte apoptosis was evenly distributed throughout the white matter whereas neuroapoptosis occurred primarily in the cortex (all regions), caudate, putamen and thalamus. Conclusions A 3 h exposure to ISO is sufficient to induce widespread neurotoxicity in the developing primate brain. These results are relevant for clinical medicine, as many surgical and diagnostic procedures in children require anaesthesia durations similar to those modelled here. Further research is necessary to identify long-term neurobehavioural consequences of 3 h ISO exposure.

Medical termination of pregnancy in cynomolgus macaques

Although pregnancy is expected during studies of novel contraceptives in non‐human primates, gestation, delivery, and lactation remove females from groups for prolonged intervals. As the macaque cervix does not facilitate transcervical surgical termination of pregnancy, we sought to establish a medical termination protocol.

The human IL-15 superagonist ALT-803 directs SIV-specific CD8+ T cells into B-cell follicles.

Sequestering of latent HIV in follicular helper T cells within B-cell follicles that largely exclude cytotoxic T cells is a major barrier to cellular immune-based approaches to eradicate HIV. Here, we show that the clinical-grade human interleukin-15 (IL-15) superagonist ALT-803 activates and redirects simian immunodeficiency virus (SIV)-specific CD8+ T cells from the peripheral blood into B-cell follicles. In agreement with the increased trafficking of SIV-specific cytotoxic T cells to sites of cryptic viral replication, lymph nodes of elite controlling macaques contained fewer cells expressing SIV RNA or harboring SIV DNA post-ALT-803 treatment. These data establish ALT-803 as an immunotherapeutic for HIV and other chronic viral pathogens that evade host immunity by persisting in B-cell follicles.

Allogeneic stem cell transplantation in fully MHC-matched Mauritian cynomolgus macaques recapitulates diverse human clinical outcomes

Allogeneic hematopoietic stem cell transplantation (HSCT) is a critically important therapy for hematological malignancies, inborn errors of metabolism, and immunodeficiency disorders, yet complications such as graft-vs.-host disease (GvHD) limit survival. Development of anti-GvHD therapies that do not adversely affect susceptibility to infection or graft-vs.-tumor immunity are hampered by the lack of a physiologically relevant, preclinical model of allogeneic HSCT. Here we show a spectrum of diverse clinical HSCT outcomes including primary and secondary graft failure, lethal GvHD, and stable, disease-free full donor engraftment using reduced intensity conditioning and mobilized peripheral blood HSCT in unrelated, fully MHC-matched Mauritian-origin cynomolgus macaques. Anti-GvHD prophylaxis of tacrolimus, post-transplant cyclophosphamide, and CD28 blockade induces multi-lineage, full donor chimerism and recipient-specific tolerance while maintaining pathogen-specific immunity. These results establish a new preclinical allogeneic HSCT model for evaluation of GvHD prophylaxis and next-generation HSCT-mediated therapies for solid organ tolerance, cure of non-malignant hematological disease, and HIV reservoir clearance.Rhesus macaques are not ideal for studying response to allogeneic hematopoietic stem cell transplant (allo-HSCT) owing to complex MHC genetics that prevent full MHC-matching. Here the authors show that inbred Mauritian-origin cynomolgus macaques are a superior preclinical model of allogeneic stem cell transplantation that mimics diverse clinical outcomes of human allo-HSCT.

Caffeine Augments Anesthesia Neurotoxicity in the Fetal Macaque Brain

Caffeine is the most frequently used medication in premature infants. It is the respiratory stimulant of choice for apnea associated with prematurity and has been called the silver bullet in neonatology because of many proven benefits and few known risks. Research has revealed that sedative/anesthetic drugs trigger apoptotic death of neurons and oligodendrocytes in developing mammalian brains. Here we evaluated the influence of caffeine on the neurotoxicity of anesthesia in developing nonhuman primate brains. Fetal macaques (n = 7–8/group), at a neurodevelopmental age comparable to premature human infants, were exposed in utero for 5 hours to no drug (control), isoflurane, or isoflurane + caffeine and examined for evidence of apoptosis. Isoflurane exposure increased apoptosis 3.3 fold for neurons and 3.4 fold for oligodendrocytes compared to control brains. Isoflurane + caffeine caused neuronal apoptosis to increase 8.0 fold compared to control levels but did not augment oligoapoptosis. Neuronal death was particularly pronounced in the basal ganglia and cerebellum. Higher blood levels of caffeine within the range considered therapeutic and safe for human infants correlated with increased neuroapoptosis. Caffeine markedly augments neurotoxicity of isoflurane in the fetal macaque brain and challenges the assumption that caffeine is safe for premature infants.