Study of the Expression Transition of Cardiac Myosin Using Polarization-Dependent SHG Microscopy

Abstract

Detection of the transition between the two myosin isoforms aand b-myosin in living cardiomyocytes is essential for understanding cardiac physiology and pathology. In this study, the differences in symmetry of polarization spectra obtained from aand b-myosin in various mammalian ventricles and propylthiouracil-treated rats are explored through polarization-dependent second harmonic generation microscopy. Here, we report for the, to our knowledge, first time that aand b-myosin, as protein crystals, possess different symmetries: the former has C6 symmetry, and the latter has C3v. A single-sarcomere line scan further demonstrated that the differences in polarization-spectrum symmetry between aand b-myosin came from their head regions: the head and neck domains of aand b-myosin account for the differences in symmetry. In addition, the dynamic transition of the polarization spectrum from C6 to C3v line profile was observed in a cell culture in which norepinephrine induced an ato b-myosin transition.

Monitoring aging‐associated structural alterations in Caenorhabditis elegans striated muscles via polarization‐dependent second‐harmonic generation measurements

“…Sarcomeres of C. elegans exhibit progressive F I G U R E 6 A, Mean DIF computed with DFT based on Equation ( 8) and estimated with a fitting procedure based on trigonal symmetry's Equations ( 9), ( 17) and ( 18) for the same samples as disorganization, with less tight packing and some disorientation of myosin molecules with aging [3,4]. In addition, a very recent study [17] has observed that there is a transition of SHG modulation with respect to the laser polarization in rat ventricles myosin filaments from a cylindrical to trigonal symmetry line profile during aging. The obtained results are in accordance with our study.…”

“…Studies have used PSHG measurements for the functional imaging of muscle cells and the in-vivo monitoring of conformational changes in myosin [15,16]. Moreover, structural modifications of cardiac myosin filaments have been detected, and the discrimination of a-and b-myosin in cardiac samples via PSHG was possible [17,18]. In the current work, we used PSHG imaging microscopy to measure aging-related myosin structural modifications in the striated muscles of C. elegans samples.…”

Towards super‐resolved terahertz microscopy for cellular imaging

“…NIR imaging is currently considered a well‐integrated option in intravital microscopy, since it can be easily deployed on point‐scanning laser systems, in conjunction with visible range laser lines that are adopted for confocal microscopy. The advantage of using NIR tuneable pulsed lasers relies on the possibility of visualising in‐depth fluorescent tissues 22 and to obtain, as a label‐free signal, the second harmonic generation (SHG) from biologically relevant non‐centrosymmetric structures such as collagen 23 and myosin 24 . Furthermore, in multiphoton microscopy, the excitation probability decreases very rapidly as function of the distance from the focal plane, conferring then an intrinsic spatial confinement to the generated fluorescence or label‐free (SHG) signal; indeed, the effective excitation volume is in the order of a femtolitre 25 …”

Stokes polarimetry-based second harmonic generation microscopy for collagen and skeletal muscle fiber characterization

“…SHG image contrast is proportional to the molecular structure and order of the myosin. Also, Yuan et al demonstrated that α- and β-myosin possess different symmetries that were observed under polarization-resolved SHG microscopy in a single-sarcomere line scan [ 42 ]. Again, the changes in the structure and physiological properties of myofibrils are investigated through the intrinsic SH signal [ 5 , 37 ].…”