Amelioration of Penetrating Ballistic-Like Brain Injury Induced Cognitive Deficits after Neuronal Differentiation of Transplanted Human Neural Stem Cells

Penetrating traumatic brain injury (PTBI) is one of the major cause of death and disability worldwide. Previous studies with penetrating ballistic-like brain injury (PBBI), a PTBI rat model revealed widespread perilesional neurodegeneration, similar to that seen in humans following gunshot wound to the head, which is unmitigated by any available therapies to date. Therefore, we evaluated human neural stem cell (hNSC) engraftment to putatively exploit the potential of cell therapy that has been seen in other central nervous system injury models. Toward this objective, green fluorescent protein (GFP) labeled hNSC (400,000 per animal) were transplanted in immunosuppressed Sprague–Dawley (SD), Fisher, and athymic (ATN) PBBI rats 1 week after injury. Tacrolimus (3 mg/kg 2 days prior to transplantation, then 1 mg/kg/day), methylprednisolone (10 mg/kg on the day of transplant, 1 mg/kg/week thereafter), and mycophenolate mofetil (30 mg/kg/day) for 7 days following transplantation were used to confer immunosuppression. Engraftment in SD and ATN was comparable at 8 weeks post-transplantation. Evaluation of hNSC differentiation and distribution revealed increased neuronal differentiation of transplanted cells with time. At 16 weeks post-transplantation, neither cell proliferation nor glial lineage markers were detected. Transplanted cell morphology was similar to that of neighboring host neurons, and there was relatively little migration of cells from the peritransplant site. By 16 weeks, GFP-positive processes extended both rostrocaudally and bilaterally into parenchyma, spreading along host white matter tracts, traversing the internal capsule, and extending ∼13 mm caudally from transplantation site reaching into the brainstem. In a Morris water maze test at 8 weeks post-transplantation, animals with transplants had shorter latency to platform than vehicle-treated animals. However, weak injury-induced cognitive deficits in the control group at the delayed time point confounded benefits of durable engraftment and neuronal differentiation. Therefore, these results justify further studies to progress towards clinical translation of hNSC therapy for PTBI

Sociosexual and Communication Deficits after Traumatic Injury to the Developing Murine Brain

<div><p>Despite the life-long implications of social and communication dysfunction after pediatric traumatic brain injury, there is a poor understanding of these deficits in terms of their developmental trajectory and underlying mechanisms. In a well-characterized murine model of pediatric brain injury, we recently demonstrated that pronounced deficits in social interactions emerge across maturation to adulthood after injury at postnatal day (p) 21, approximating a toddler-aged child. Extending these findings, we here hypothesized that these social deficits are dependent upon brain maturation at the time of injury, and coincide with abnormal sociosexual behaviors and communication. Age-dependent vulnerability of the developing brain to social deficits was addressed by comparing behavioral and neuroanatomical outcomes in mice injured at either a pediatric age (p21) or during adolescence (p35). Sociosexual behaviors including social investigation and mounting were evaluated in a resident-intruder paradigm at adulthood. These outcomes were complemented by assays of urine scent marking and ultrasonic vocalizations as indices of social communication. We provide evidence of sociosexual deficits after brain injury at p21, which manifest as reduced mounting behavior and scent marking towards an unfamiliar female at adulthood. In contrast, with the exception of the loss of social recognition in a three-chamber social approach task, mice that received TBI at adolescence were remarkably resilient to social deficits at adulthood. Increased emission of ultrasonic vocalizations (USVs) as well as preferential emission of high frequency USVs after injury was dependent upon both the stimulus and prior social experience. Contrary to the hypothesis that changes in white matter volume may underlie social dysfunction, injury at both p21 and p35 resulted in a similar degree of atrophy of the corpus callosum by adulthood. However, loss of hippocampal tissue was greater after p21 compared to p35 injury, suggesting that a longer period of lesion progression or differences in the kinetics of secondary pathogenesis after p21 injury may contribute to observed behavioral differences. Together, these findings indicate vulnerability of the developing brain to social dysfunction, and suggest that a younger age-at-insult results in poorer social and sociosexual outcomes.</p></div

3chamber_test_data

Mice injured at adolescence (postnatal day 35) and tested in the three-chamber social approach task at adulthood. Percentage time spent in each chamber was quantified

BodyWeights_data

Body weights of mice used in this study, across time post-injur

Res-Intr_test_data

Social investigation of a novel (male or female) stimulus mouse was quantified as duration spent investigating different regions (e.g. anogenital), during the resident-intruder task

CC_volume_data

Stereological analyses were performed using StereoInvestigator to determine the volume loss of the corpus callosum at adulthood after injury at either p21 or p35

Scent_mark_test_data

Grid squares containing scent marks were quantified during the scent marking test towards a novel female mouse, of sham or TBI (traumatic brain injury) mice at adulthood after injury at p21 or p35

USV_data

Ultrasonic vocalizations (USVs) were recorded during encounters of the test mouse (sham or brain-injured) with (1) a novel male, (2) a novel female, or (3) soiled bedding from a novel female. Several parameters including total number of calls, high frequency calls (> 75 kHz) and # calls in bursts were quantified

Reduced social investigation at adulthood after pediatric but not adolescent TBI.

<p>Experimental timelines illustrate the timing of behavioral assessments; Cohort 1 (a) consisted of n = 10/group after TBI or sham-operation at p21; Cohort 2 (b) was n = 7–8/group, also after TBI or sham at p21; Cohort 3 (c) consisted of mice that received TBI or sham-operation at p35 (adolescence; n = 9/group). Investigative and interactive behaviors by sham and TBI mice, at adulthood after p21 or p35 injury, were quantified after addition of a novel male mouse in the resident-intruder (RI) paradigm (d). Brain-injured mice that received TBI at p21 (e) spent less time engaged in social investigative behaviors including ano-genital sniffing and following (**p<0.01, *p<0.05). Antagonistic behaviors (f), quantified as the number of fighting bouts and latency to first fight, were not different between the injury groups. After injury at p35 (g, h), sham and TBI mice also showed similar investigative behaviors towards a novel male mouse.</p