Formalin fixation on HER-2 and PD-L1 expression in gastric cancer:A pilot analysis using the same surgical specimens with different fixation times

BACKGROUND The needs for human epidermal growth factor receptor 2(HER-2) and/or programmed death-ligand 1(PD-L1) evaluations in gastric cancer are dramatically increasing. Although the importance of standardization of sample fixation has been widely recognized, most of the evidence regarding the fixation duration or type of fixing solution are based on breast cancer.AIM To investigate the real effects of fixation conditions on HER-2 testing or PD-L1 testing for gastric cancer using gastrectomy specimens.METHODS Thirty-two patients who underwent gastrectomy for gastric cancer were enrolled.Their resected specimens were each divided into four pieces and fixed in four strictly controlled different durations(6 h, 24 h, and 48 h, and 1 wk) by 10%formalin(n = 22) or 10% neutral buffered formalin(NBF)(n = 10).Immunohistochemistry(IHC) of HER-2 and PD-1 was performed, and a pathology examination was conducted. In the HER-2-immunoreactive cases, all four specimens were subjected to dual-color in situ hybridization(DISH). Five cases were assessed as HER-2-positive by IHC and DISH. We used the cut-off values of 1%, 10%, and 50% to assess the IHC findings of PD-L1.RESULTS No significant difference was observed in comparisons between the shorter fixation period groups(6 h, 24 h, and 48 h) and the prolonged fixation period(1 wk) group in the HER-2 and PD-L1 analyses. Although no significant difference was observed between 10% formalin and 10% NBF within 1 wk of fixation, the superiority of 10% NBF was confirmed in a long-term(> 3 mo) fixation in both the HER-2 and PD-L1 analyses.CONCLUSION In this small-numbered pilot study, prolonged fixation within 1 wk showed no inferiority in HER-2 or PD-L1 testing. However, a large-numbered prospective study is needed to obtain conclusive results.

Characterization of Rebound Depolarization in Neurons of the Rat Medial Geniculate Body In Vitro

Rebound depolarization （RD） is a response to the offset from hyperpolarization of the neuronal mem- brane potential and is an important mechanism for the synaptic processing of inhibitory signals. In the present study, we characterized RD in neurons of the rat medial geniculate body （MGB）, a nucleus of the auditory thala- mus, using whole-cell patch-clamp and brain slices. RD was proportional in strength to the duration and magnitude of the hyperpolarization; was effectively blocked by Ni2＋ or Mibefradil; and was depressed when the resting membrane potential was hyperpolarized by blocking hyperpolarization-activated cyclic nucleotide-gated （HCN） channels with ZD7288 or by activating G-protein-gated inwardly-rectifying K＋ （GIRK） channels with baclofen. Our results demonstrated that RD in MGB neurons, which is carried by T-type Ca2＋ channels, is critically regulated by HCN channels and likely by GIRK channels.