Evaluation of in vivo bioactivities of recombinant hypo- (FSH21/18) and fully- (FSH24) glycosylated human FSH glycoforms in Fshb null mice

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Wang, Huizhen
May, Jacob
Butnev, Viktor Y.
Shuai, Bin
May, Jeffrey V.
Bousfield, George R.
Kumar, T. Rajendra
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FSH-responsive genes , N-glycosylation , Ovary , Testis , Pharmacological rescue
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Wang, Huizhen; May, Jacob; Butnev, Viktor Y.; Shuai, Bin; May, Jeffrey V.; Bousfield, George R.; Kumar, T. Rajendra. 2016. Evaluation of in vivo bioactivities of recombinant hypo- (FSH21/18) and fully- (FSH24) glycosylated human FSH glycoforms in Fshb null mice. Molecular and Cellular Endocrinology, vol. 437, 5 December 2016:pp 224–236

The hormone - specific FSH beta subunit of the human FSH heterodimer consists of N-linked glycans at Asn(7) and Asn(24) residues that are co-translationally attached early during subunit biosynthesis. Differences in the number of N-glycans (none, one or two) on the human FSH beta subunit contribute to macroheterogeneity in the FSH heterodimer. The resulting FSH glycoforms are termed hypo-glycosylated (FSH21/18, missing either an Asn(24) or Asn(7) N-glycan chain on the beta - subunit, respectively) or fully glycosylated (FSH24, possessing of both Asn(7) and Asn(24) N-linked glycans on the beta-subunit) FSH. The recombinant versions of human FSH glycoforms (FSH21/18 and FSH24) have been purified and biochemically characterized. In vitro functional studies have indicated that FSH21/18 exhibits faster FSH- receptor binding kinetics and is much more active than FSH24 in every assay tested to date. However, the in vivo bioactivity of the hypo-glycosylated FSH glycoform has never been tested. Here, we evaluated the in vivo bioactivities of FSH glycoforms in Fshb null mice using a pharmacological rescue approach. In Fshb null female mice, both hypo- and fully-glycosylated FSH elicited an ovarian weight gain response by 48 h and induced ovarian genes in a dose- and time-dependent manner. Quantification by real time qPCR assays indicated that hypo-glycosylated FSH21/18 was bioactive in vivo and induced FSH-responsive ovarian genes similar to fully-glycosylated FSH24. Western blot analyses followed by densitometry of key signaling components downstream of the FSH-receptor confirmed that the hypo-glycosylated FSH21/18 elicited a response similar to that by fully-glycosylated FSH24 in ovaries of Fshb null mice. When injected into Fshb null males, hypo-glycosylated FSH21/18 was more active than the fully-glycosylated FSH24 in inducing FSH-responsive genes and Sertoli cell proliferation. Thus, our data establish that recombinant hypo-glycosylated human FSH21/18 glycoform elicits bioactivity in vivo similar to the fully-glycosylated FSH. Our studies may have clinical implications particularly in formulating FSH-based ovarian follicle induction protocols using a combination of different human FSH glycoforms.

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Elsevier, Ireland LTD
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Molecular and Cellular Endocrinology;v.437
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