Supplementation of in vitro culture medium with FSH to grow follicles and mature oocytes can be replaced by extracts of Justicia insularis

PLoS One. 2018 Dec 7;13(12):e0208760. doi: 10.1371/journal.pone.0208760. eCollection 2018.

Abstract

The present study evaluated the effect of supplementing in vitro culture medium with J. insularis compared to FSH on isolated secondary follicles and in vitro maturation of oocytes from those follicles. Secondary follicles were isolated from sheep ovaries and individually cultured for 18 days in α-MEM+ (Control), α-MEM+ supplemented with 100 ng/mL recombinant bovine follicle stimulating hormone (FSH) or with 0.3, 1.25, or 2.5 mg/mL of J. insularis extract (JI0.3, JI1.25, and JI2.5, respectively). Culture medium collected every 2 days was used to measure ROS levels. At the end of the culture period, cumulus oocytes complex (COCs) were collected and matured in vitro. Follicular walls were used for mRNA quantitation. JI0.3 led to a higher (P < 0.05) percentages of intact follicles than other groups after 18 days of culture. While follicular diameter remained unchanged from Day 6 onwards with JI0.3 and FSH, percentages of antral cavity formation were higher (P < 0.05) with JI0.3 at Day 6 than in all other treatments. No differences were observed between controls and treatment groups regarding ROS levels and mRNA expression of genes. Viability of resulting oocytes was higher (P < 0.05) in JI0.3 compared to FSH. Interestingly, in control experiment, supplementation of maturation medium with JI0.3 led to higher (P < 0.05) percentages of metaphase II compared to controls. Although more validations will be needed, it seems that this natural extract could be used as a cheap and easily available alternative to commercial FSH.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Culture Media*
  • Dose-Response Relationship, Drug
  • Female
  • Follicle Stimulating Hormone / administration & dosage*
  • Gene Expression Regulation, Developmental
  • In Vitro Oocyte Maturation Techniques*
  • Justicia* / chemistry
  • Oocytes / drug effects
  • Oocytes / growth & development
  • Oocytes / metabolism
  • Oogenesis / drug effects
  • Oogenesis / physiology
  • Ovarian Follicle / drug effects
  • Ovarian Follicle / growth & development
  • Ovarian Follicle / metabolism
  • Phytochemicals / administration & dosage
  • Phytochemicals / chemistry
  • Plant Extracts / administration & dosage*
  • Plant Extracts / chemistry
  • Plant Leaves / chemistry
  • RNA, Messenger / metabolism
  • Reactive Oxygen Species / metabolism
  • Sheep

Substances

  • Culture Media
  • Phytochemicals
  • Plant Extracts
  • RNA, Messenger
  • Reactive Oxygen Species
  • Follicle Stimulating Hormone

Grants and funding

This research was supported by grants from the National Council for Scientific and Technological Development (CNPq Brazil) for financial support. Gildas Tetaping Mbemya is the recipient of a doctoral scholarship from Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (PEC-PG/CAPES, Brazil). Johan Smitz is Especial Visitor Researcher from Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES: 88881.030.433/2013-01) and Ana Paula Ribeiro Rodrigues is recipient of a grant from CNPq Brazil through the projects 473968/2013-4 and 457226/2013-7.