Different types of Retinoblastoma seeds with high risk factors: IJO

Das D, Bhattacharjee K, Barman MJ, Bhattacharjee H, Maity S, Bandyopadhyay D, et al. studied the pathological significance of RB seeds with histopathological risk factors. Field emission scanning electron microscopy (FESEM) for seeds, calcifications and Raman spectroscopy were studied in an index case in the cohort with interesting observations.

This study was a laboratory‑based observational study. Enucleated eyeballs received in the ocular pathology department of a tertiary eye care center in northeast India were included in the cohort after obtaining written informed consent during the surgery. The study was carried out for 6 years (2015–2020). Most of the eyeballs were Group‑E RBs. Standard eyeballs sectioning were done by bread loaf techniques. Gross documentations included RB seeds seen in the smallest calotte done with utmost care. Seeds were documented also in permanent sections. Scanning electron microscopy and Raman spectroscopy were carried out in an index case.

• Out of the total 59 cases, 35 RB cases had different seedings. The mean age at enucleation was 2.9 years.

• RB seeds were seen in vitreous (n = 19), subretinal plus vitreous (n = 7), anterior chamber (n = 1), over crystalline lens (n = 3), retinal surface (n = 1), retinal pigment epithelium (RPE; n = 2), subretinal (n = 1), calcified seeds (n = 2).

• Other characteristics were dusts (n = 7), clouds (n = 11), spheres (n = 4), and unspecified type (n = 13). Histopathological high‑risk factors showed significant choroidal (n = 22) and optic nerve (n = 15) involvement.

• Few cases had extraocular spread. Undifferentiated tumor (n = 24) was seen with higher evidence of necrosis (n = 23).

• Raman spectra differentiated the seeds from the normal tissue on the basis of lipid and protein content.

RB is the most common primary intraocular malignancy in childhood. It is a cancer of the retina that is seen as an extension of the brain to the eye. This pathological study at a tertiary institute revealed 35 cases of RB with different seeds. The most common seeds were located in the vitreous cavity. Isolated seeds were seen in the anterior chamber, which was depository type, retinal, and subretinal tissue.

The surface of vitreous RB seeds differed from subretinal ones in study. RB seeds were correlated with other histopathological findings. Endophytic growth was the maximum in this study. High‑risk histopathological factors showed significant choroidal involvements in lateral calottes of eyeball along with optic nerve involvement. The active seeds in the vitreous, subretinal, and other extraocular spaces were associated with the advancement of the cancerous growth. Undifferentiated tumors were more than the differentiated ones. Tumor necrosis and anaplasia were noted in a significant number of cases. Authors had seen anaplasia in 17 cases. Most of the anaplasias were graded as moderate to severe (15/17 cases).

The present study included FESEM observations of RB seeds, intratumoral calcification, and Raman spectroscopy study in a single case. This was in an enucleated eyeball in an 18‑month‑old boy with a Group‑ERB, where direct visualization of vitreous and subretinal seeds was made under fluorescein stain and compared with the seeds in H and E‑stained slide and FESEM. There was full‑thickness choroidal involvement in that eyeball with prelaminar optic nerve involvement. The tumor was undifferentiated with extensive necrosis and calcification. The surface pigment change of subretinal seeds and vitreous seeds having honeycomb appearance correlated with the H and E‑stained slides' seeds. FESEM‑documented seeds could suggest that those surface pigments on subretinal seeds could have local effects in tumor resistance to chemotherapy. RB seed documentation in FESEM has never been reported in the scientific literature previously. RB calcification in the same case by FESEM showed a lamellar arrangement of osteophytes with fiber orientations that changed from one lamella to the adjacent. The remodeling of tumoral calcification observation in RB calcification was also not documented by FESEM previously in the literature, as in the case in the study.

Noninfrared Raman spectroscopy in study showed important findings between normal adjoining tissue and a group of RB seeds. Spectra of normal tissue showed less spectral spikes, whereas RB seeds showed characteristic high spikes. This can be attributed to normal tissue that has more lipids content, whereas malignant RB seeds havehigher protein by‑products expressed by the tumor. Raman spectroscopy can map useful cellular chemico‑physical microenvironment of the tumor.

This pathological study on RB seeds showed various high‑risk factors associated with the tumor seeds. Characteristic surface change of these subretinal seeds could be associated with RB chemotherapy resistances that require future validation. FESEM and Raman spectroscopy in an index case RB seeds has opened up a new frontier to ocular tumor imaging in the future.