Supplementary Material for: Recurrent Moderate Hypoglycemia Suppresses Brain-Derived Neurotrophic Factor Expression in the Prefrontal Cortex and Impairs Sensorimotor Gating in the Posthypoglycemic Period in Young Rats

Recurrent hypoglycemia is common in infants and children. In developing rat models, recurrent moderate hypoglycemia leads to neuronal injury in the medial prefrontal cortex. To understand the effects beyond neuronal injury, 3-week-old male rats were subjected to 5 episodes of moderate hypoglycemia (blood glucose concentration, approx. 30 mg/dl for 90 min) once daily from postnatal day 24 to 28. Neuronal injury was determined using Fluoro-Jade B histochemistry on postnatal day 29. The effects on brain-derived neurotrophic factor (BDNF) and its cognate receptor, tyrosine kinase receptor B (TrkB) expression, which is critical for prefrontal cortex development, were determined on postnatal day 29 and at adulthood. The effects on prefrontal cortex-mediated function were determined by assessing the prepulse inhibition of the acoustic startle reflex on postnatal day 29 and 2 weeks later, and by testing for fear-potentiated startle at adulthood. Recurrent hypoglycemia led to neuronal injury confined primarily to the medial prefrontal cortex. BDNF/TrkB expression in the prefrontal cortex was suppressed on postnatal day 29 and was accompanied by lower prepulse inhibition, suggesting impaired sensorimotor gating. Following the cessation of recurrent hypoglycemia, the prepulse inhibition had recovered at 2 weeks. BDNF/TrkB expression in the prefrontal cortex had normalized and fear-potentiated startle was intact at adulthood. Recurrent moderate hypoglycemia during development has significant adverse effects on the prefrontal cortex in the posthypoglycemic period

The CD4 receptor plays essential but distinct roles in HIV-1 infection and induction of apoptosis in primary bone marrow GPIIb/IIIa +

We investigated whether cells belonging to the megakaryocytic lineage could be infected in vitro with human immunodeficiency virus type-1 (HIV-1). Primary GPIIb/IIIa+ bone marrow (BM) cells and HEL continuous cell line were first phenotypically characterized for the presence of megakaryocytic markers and CD4 antigen, then challenged in vitro with the laboratory strain IIIB of HIV-1. Both GPIIb/IIIa+ BM and HEL cells expressed significant levels of CD4 receptor (> 50%) and were efficiently infected with HIV-1, as judged by the presence of proviral DNA after polymerase chain reaction analysis and by quantitative evaluation of gag p24 antigen in the culture supernatants. Of note, infection with HIV-1 in both primary BM megakaryocytes and HEL cells was specifically blocked by soluble recombinant CD4. To ascertain whether the CD4 receptor was essential for infection of megakaryocytic cells, HEL were subcloned into CD4+ and CD4- cells. Although unfractionated and CD4+ HEL cells were productively infected with HIV-1, CD4- HEL cells could not be infected. Infection of HEL cells did not induce gross cytotoxic effects or a significant increase of apoptosis. On the other hand, treatment of unfractionated or CD4+ HEL cells with cross-linked recombinant env gp120 or Leu3a anti-CD4 monoclonal antibody markedly (P < 0.01) increased the degree of apoptosis with respect to HEL cells infected with HIV-1 or treated with cross-linked gag p24 or anti-GPIIb/IIIa antibody. Taken together, these data indicate that the CD4 receptor represents the main route of infection in cells belonging to the megakaryocytic lineage. Moreover, an inappropriate engagement of CD4 by either free env gp120 or anti-CD4 monoclonal antibody could be more relevant than a direct infection with HIV-1 in the induction of the frequent BM megakaryocyte abnormalities found in HIV-1 seropositive thrombocytopenic patients