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Stanley Ka-lok Li, Juni Banerjee, Christopher Jang, Amita Sehgal, Richard A Stone, Mortimer Civan; Physiological Temperature Oscillations Alter Matrix Metalloproteinase MMP-2,9 Secretion by TM5 Human Trabecular Meshwork Cells Independent of Clock Genes Per2 and Cry2. Invest. Ophthalmol. Vis. Sci. 2014;55(13):2903.
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© ARVO (1962-2015); The Authors (2016-present)
Aqueous humor inflow falls by 50% during sleeping hours but is unaccompanied by consistent change in intraocular pressure, partly because of reduced outflow facility (OF). One regulator of OF is MMP release from trabecular meshwork (TM) cells. We tested if OF cycling might result from circadian cycling of peripheral clock genes triggered by changes in core temperature initiated by the suprachiasmatic nucleus central clock and by closing eyelids in sleep.
Transformed normal human TM5 cells were provided by Alcon/Novartis. Temperature was constant or underwent 12hr/12hr cycling between 33 and 37 deg C. Diurnal MMP-2,9 secretion was measured by zymography and gene expression of Per2 and Cry2 clock genes by real-time PCR (qPCR).
Raising temperature to 37 deg C increased, and lowering to 33 reduced, MMP release. Step changes in temperature (n=494, 5 experiments) altered MMP-9 release by 46+/-3% and MMP-2 by 22+/-2%. However, even after 9 days of temperature cycling, oscillations were not later maintained at constant temperature. Inhibition of heat shock transcription factor 1 (HSF1) with KNK437 (100 μM) inhibited MMP-9, but little affected MMP-2 release during temperature cycling. Transient activation of Per2 and Cry2 was observed at 37 deg C, but temperature cycling did not further change expression.
Alternating temperature over a physiologic range alters MMP-9 more than MMP-2 release, consistent with temperature-driven cycling of peripheral circadian clocks and perhaps daily cycling of OF. However, the absence of entrainment and limited inhibition of cycling of MMP-2 release produced by blocking HSF1 suggest non-circadian mediation. The clock genes Per2 and Cry2 also did not appear to cycle in response to temperature in these cells. If cycling of OF reflects a TM-cell circadian rhythm, the triggering stimulus to the TM clock genes is not temperature, and regulators other than MMP-2,9 may be the mediators.
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