Utility of microtubule associated protein-2 (MAP-2) immunohistochemistry for identification of ganglion cells in paraffin-embedded rectal suction biopsies.

Identification of rare ganglion cells in rectal suction biopsies can be difficult for surgical pathologists who only occasionally encounter rectal suction biopsies for the diagnosis of Hirschsprung disease. We evaluated the utility of a monoclonal antibody directed against microtubule associated protein-2 (MAP-2), a marker of neuronal differentiation, to detect ganglion cells in formalin-fixed, paraffin-embedded tissue sections. The presence of ganglion cells was evaluated retrospectively by immunohistochemical staining for MAP-2 in 78 biopsies from 56 patients with ganglion cells present on hematoxylin and eosin (H&E) stain and 91 biopsies from 46 patients with no ganglion cells present on H&E stain. A total of 78 biopsies from 49 patients diagnosed as equivocal or suboptimal for ganglion cells on H&E stain were also examined. MAP-2 antibody clearly highlighted ganglion cell bodies in all 78 biopsies with ganglion cells present on H&E stain, including immature forms, and was negative in 88 biopsies with no ganglion cells present on H&E stain. In 3 biopsies from 2 patients with no ganglion cells present on H&E stain, MAP-2 antibody highlighted rare ganglion cells and in each case, the patient was confirmed not to have Hirschsprung disease on follow-up biopsy. MAP-2 immunoreactive cells were also identified in 14 biopsies from 12 patients diagnosed as equivocal for identification of ganglion cells on H&E stain. Three of these patients were found not to have Hirschsprung disease on repeat rectal suction biopsy and 6 additional patients in this group showed no evidence of Hirschsprung disease by clinical follow-up. This study indicates that MAP-2 antibody is a sensitive and specific immunohistochemical marker that can confirm ganglion cells in paraffin-embedded rectal suction biopsies and may eliminate the need for repeat biopsy in select cases.

How we treat: management of life-threatening primary postpartum hemorrhage with a standardized massive transfusion protocol.

Management of massive, life-threatening primary postpartum hemorrhage in the labor and delivery service is a challenge for the clinical team and hospital transfusion service. Because severe postpartum obstetrical hemorrhage is uncommon, its occurrence can result in emergent but variable and nonstandard requests for blood products. The implementation of a standardized massive transfusion protocol for the labor and delivery department at our institution after a maternal death caused by amniotic fluid embolism is described. This guideline was modeled on a existing protocol used by the trauma service mandating emergency release of 6 units of group O D- red cells (RBCs), 4 units of fresh frozen or liquid plasma, and 1 apheresis unit of platelets (PLTs). The 6:4:1 fixed ratio of uncrossmatched RBCs, plasma, and PLTs allows the transfusion service to quickly provide blood products during the acute phase of resuscitation and allows the clinical team to anticipate and prevent dilutional coagulopathy. The successful management of three cases of massive primary postpartum hemorrhage after the implementation of our new massive transfusion protocol in the maternal and fetal medicine service is described.

Genotoxin-induced Rad9-Hus1-Rad1 (9-1-1) chromatin association is an early checkpoint signaling event.

Rad17, Rad1, Hus1, and Rad9 are key participants in checkpoint signaling pathways that block cell cycle progression in response to genotoxins. Biochemical and molecular modeling data predict that Rad9, Hus1, and Rad1 form a heterotrimeric complex, dubbed 9-1-1, which is loaded onto chromatin by a complex of Rad17 and the four small replication factor C (RFC) subunits (Rad17-RFC) in response to DNA damage. It is unclear what checkpoint proteins or checkpoint signaling events regulate the association of the 9-1-1 complex with DNA. Here we show that genotoxin-induced chromatin binding of 9-1-1 does not require the Rad9-inducible phosphorylation site (Ser-272). Although we found that Rad9 undergoes an additional phosphatidylinositol 3-kinase-related kinase (PIKK)-dependent posttranslational modification, we also show that genotoxin-triggered 9-1-1 chromatin binding does not depend on the catalytic activity of the PIKKs ataxia telangiectasia-mutated (ATM), ataxia telangiectasia and Rad3-related (ATR), or DNA-PK. Additionally, 9-1-1 chromatin binding does not require DNA replication, suggesting that the complex can be loaded onto DNA in response to DNA structures other than stalled DNA replication forks. Collectively, these studies demonstrate that 9-1-1 chromatin binding is a proximal event in the checkpoint signaling cascade.