Patients
Drug resistant MTLE-HS patients (n = 55) underwent standard presurgical
workups which include video electroencephalogram (vEEG), epilepsy
protocol MRI, interictal fluoro-2-deoxyglucose positron emission
tomography (FDG-PET), ictal single photon emissioncomputer tomography
(SPECT) and magnetoencephalography (MEG) [22]. Patient details were
discussed weekly at epilepsy surgery meeting attended by
epileptologists, epilepsysurgeon, neuroradiologists and nuclear-medicine
specialists where patients were selected for surgery based on
concordance of above-mentioned diagnostic procedures. Surgical resection
involves anterior temporal lobectomy with amygdalohippocampectomy. A
portion of resected hippocampus was analysed by neuropathologist for
confirmation of the diagnosis of MTLE-HS. All these patients were
seizure-free post-operatively (class I Engel outcome) (Supplementary
Table 1) (Engel, 1983). Resected hippocampal tissues from patients who
underwent autopsies and devoid of any history of brain pathology were
collected within 12h of death and used as autopsy controls (n= 15)
(Supplementary Table 2). For experiments involving cellular
electrophysiological studies resected cortical tissues obtained from
surrounding areas of brain tumours during surgical resection, which was
also a part of planned surgical resection, in patients with glioma
without any history of seizures were considered as non-seizure controls
(n = 20) molecular as autopsy control samples were not suitable for
these experiments (Supplementary Table 3) (Banerjee et al., 2015; 2017;
2020). Samples were collected from 2015 to 2019 at the All India
Institute of Medical Sciences in New Delhi, which is the referral center
for epilepsy surgery in northern India. All procedures performed in
studies involving human participants were in accordance with the ethical
standards of the institutional and/or national research committee and
with the 1964 Helsinki Declaration and its later amendments or
comparable ethical standards.
The study was ethically approved by the Institutional Ethics Committee,
All India Institute of Medical Sciences, New Delhi, India
(IECPG/-38/27.11.2015, RT-4/30.12.2015). Informed consent was obtained
from all patients or their legally authorized representatives prior to
inclusion in this study.
A small portion of resected tissue was stored for fixation in 0.1M
phosphate buffer containing 4% paraformaldehyde, pH 7.4 for 72 hours.
Post fixation, the tissues were dehydrated, embedded and cut into 6µM
thick sections with a microtome and collected on poly L-lysine coated
slides. Haematoxylin-eosin stained sections were used for
cytoarchitecture. Immunohistochemistry was performed using with
anti-GFAP (1:1000; Cat# ab7260, RRID: AB_305808), or with anti-NeuN
(1:1000; Cat# ab104225, RRID: AB_10711153) primary antibodies.
Secondary antibody was Biotinylated Goat Anti-Rabbit IgG antibody
(1:1000; Vector Laboratories Cat# BA-1000, RRID: AB_2313606)