Salivary Gland Protein Expression after Bion-M1 and Space Shuttle STS-135 Missions

Mednieks, Maija; Khatri, Aditi; Hand, Arthur R.

doi:10.2478/gsr-2015-0001

Figures & Tables

Cyclic AMP-PKA signaling pathways of regulated exocytosis in cell compartments. Schematic representation of cell surface beta-adrenergic receptor stimulation, signal transduction, cyclic AMP second messenger cytoplasmic and nuclear response. βAR, beta-adrenergic receptor; Gs, heterotrimeric G protein; AC, adenylate cyclase; PDE, phosphodiesterase; AKAP, A-kinase anchor protein; RII, type II PKA regulatory subunit; C, PKA catalytic subunit; ATF1, activating transcription factor 1; CREB, cyclic AMP response element binding protein; CREM, cyclic AMP responsive element modulator. The large blue sphere represents a secretory granule along a path to fuse with the cell membrane and to empty its contents of secretory proteins into the lumen.

Ultrastructure of parotid glands of flight mice. Panel A, STS-135: Acinar cells have basally located nuclei [N (binucleate cells are common)], a prominent Golgi complex (G), and abundant apically located secretory granules (SG). Intercellular canaliculus (IC). Scale bar = 2 μm. Panel B, STS-135: Intercalated duct. The duct cells have apical vacuoles (arrows) containing material endocytosed from the lumen (L). Scale bar = 2 μm. The inset shows a similar vacuole in a striated duct cell immunogold labeled for PSP, an acinar secretory protein. Scale bar = 0.5 μm. Panel C, Bion-M1: A recently formed autophagic vacuole (AV1) containing secretory proteins labeled for PSP is present in an acinar cell. An older autophagic vacuole (AV2) contains remnants of secretory granules and other organelles, but shows little labeling. Secretory granule (SG); nucleus (N). Scale bar = 1 μm. Panel D, Bion-M1: Apoptotic acinar cells (Apop) phagocytosed by a macrophage (arrowheads). Myoepithelial cell (MEC). Scale bar = 2 μm.

Immunogold labeling of parotid acinar cell secretory granules. Panel A, STS-135: Flight, PRPs. Panel B, STS-135: AEM ground control, PRPs. Panel C, Bion-M1: Flight, amylase. Panel D, Bion-M1: Habitat ground control, amylase. Intercellular canaliculus (IC); mitochondrion (M). Scale bar = 0.5 μm.

Morphometric analysis of immunogold labeling of parotid secretory proteins. Percent change in STS-135 (light bars) and Bion-M1 (dark bars) flight mice from habitat controls.

Electrophoresis, Western blotting, and densitometry of parotid proteins. Panel A, Bion-M1 control and flight parotid soluble proteins. The lanes are molecular size marker (M), flight (F), and vivarium control (C). The two bands on the Western blot (WB) are at the mobility of RII and an RII fragment (Rfr), respectively. Panel B, Densitometry of the banding pattern profiles. Vivarium control (light gray curve) superimposed with the flight (darker curve) protein electrophoretic pattern. The ordinate shows integrated density (ID) values and the abscissa shows the calculated molecular size in kilo Dalton (kD) units.

Figure 6.

Densitometric analysis of polyacrylamide gel electrophoretic protein separation and anti-RII and anti-α-amylase reactivity in STS-135 mice. Panel A: Top panel, flight; middle panel, vivarium; and bottom panel, habitat parotid gland samples. The lighter curve represents the protein profile; the shaded area is the reactivity to anti-RII antibody. The ordinate axes are integrated density values of the proteins on the left hand axis, and the integrated density values for RII and the reactivity of RII fragment (Rfr) on the right hand axis. Panel B shows parotid α-amylase Western blotting. The ordinate shows integrated density, and the error bars show ± 5% error. F, flight; C1, vivarium control; C2, habitat control.

Microarray analyses of selected parotid genes. The ordinate shows the log2 values of the ratio of the means of the normalized signal intensities determined for each gene for flight vs. habitat control. Prkar2a, protein kinase, cAMP dependent regulatory, type II alpha; Akap13, A-kinase anchor protein 13; Pde4a, phosphodiesterase 4a, cAMP specific; Adrb2, adrenergic receptor, beta 2; Adcy3, adenylate cyclase 3; Amy1, amylase 1, salivary.

Ratios of RII to Total Protein of STS-135 and Bion-M1 PAGE_

		F	C1	C2
STS-135	PAGE	70.8	69.28	69.99
	RII	5.16	41.43	24.24
	Ratio	0.073	0.60	0.35
Bion-M1	PAGE	153.6	129.0	---
	RII	21.7	16.26	---
	Ratio	0.14	0.13	---

Quantitative Immunogold Labeling of STS-135 and Bion-M1 Salivary Glands (Gold Particles/µm 2 ± SEM)_

Bion-M1Parotid	Flight	Habitat
Amylase	15.3 ± 1.11∞	10.3 ± 0.69
RII	10.1 ± 0.47	11.3 ± 0.49
PSP	37.4 ± 1.17§	30.6 ± 1.76
PRP	22.5 ± 1.61	24.8 ± 2.08
DCPP	23.1 ± 0.95	25.2 ± 1.27

Microarray Analyses of Secretory Protein Gene Expression_

	STS-135		Bion-M1
Gene ^a	Log2 F/H^b	Fold Change F/H	Log2 F/H	Fold Change F/H
Amy1	0.030	1.02	0.169*	1.124
Bpifa2	0.241§	1.18	0.170*	1.13
Lpo	0.070	1.05	0.160*	1.18
Lyz1	0.317	1.25	0.898∞	1.86
Pip	0.228*	1.17	0.169*	1.13
Prpmp5	-0.161§	0.894	-0.861∞	0.550

Body Weights of STS-135 and Bion-M1 Mice_

Group	Initial Body Wt. (g) ± SEM (n)	Final Body Wt. (g) ± SEM (n)
STS-135
Flight	20.7 ± 0.40 (7)	18.4 ± 0.52 (7)
AEM Ground Control	20.7 ± 0.31 (15)	19.4 ± 0.33 (15)
Vivarium Ground Control	---	19.7 ± 0.34 (15)
Bion-M1
Flight	26.8 ± 0.55 (6)	29.4 ± 1.73 (6)
Synchronous Vivarium Ground Control	29.2 ± 0.44 (8)	28.7 ± 0.30 (8)
Asynchronous Habitat Ground Control	27.3 ± 1.15 (7)	29.7 ± 0.83 (7)
Asynchronous Vivarium Ground Control	26.7 ± 0.78 (7)	28.9 ± 1.03 (7)

Salivary Gland Protein Expression after Bion-M1 and Space Shuttle STS-135 Missions

Figures & Tables

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Ratios of RII to Total Protein of STS-135 and Bion-M1 PAGE_

Quantitative Immunogold Labeling of STS-135 and Bion-M1 Salivary Glands (Gold Particles/µm 2 ± SEM)_

Microarray Analyses of Secretory Protein Gene Expression_

Body Weights of STS-135 and Bion-M1 Mice_

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