Highly sensitive visual restoration and protection via ectopic expression of chimeric rhodopsin in mice.

Photoreception requires amplification by mammalian rhodopsin through G protein activation, which requires a visual cycle. To achieve this in retinal gene therapy, we incorporated human rhodopsin cytoplasmic loops into Gloeobacter rhodopsin, thereby generating Gloeobacter and human chimeric rhodopsin (GHCR). In a murine model of inherited retinal degeneration, we induced retinal GHCR expression by intravitreal injection of a recombinant adeno-associated virus vector. Retinal explant and visual thalamus electrophysiological recordings, behavioral tests, and histological analysis showed that GHCR restored dim-environment vision and prevented the progression of retinal degeneration. Thus, GHCR may be a potent clinical tool for the treatment of retinal disorders.

Starburst amacrine cells amplify optogenetic visual restoration through gap junctions.

Ectopic induction of optogenetic actuators, such as channelrhodopsin, is a promising approach to restoring vision in the degenerating retina. However, the cell type-specific response of ectopic photoreception has not been well understood. There are limits to obtaining efficient gene expression in a specifically targeted cell population by a transgenic approach. In the present study, we established a murine model with high efficiency of gene induction to retinal ganglion cells (RGCs) and amacrine cells using an improved tetracycline transactivator-operator bipartite system (KENGE-tet system). To investigate the cell type-specific visual restorative effect, we expressed the channelrhodopsin gene into RGCs and amacrine cells using the KENGE-tet system. As a result, enhancement in the visual restorative effect was observed to RGCs and starburst amacrine cells. In conclusion, a photoresponse from amacrine cells may enhance the maintained response of RGCs and further increase or improve the visual restorative effect.

Changes in homocysteine and non-mercaptoalbumin levels after acute exercise: a crossover study.

BACKGROUND: Acute exercise is one factor that increases blood homocysteine levels, and elevated homocysteine levels cause oxidative stress. Albumin, which is abundant in blood, is an antioxidant, and the redox state of albumin is used as an index of oxidative stress in blood. This study aimed to assess the effect of acute exercise on plasma homocysteine levels and the blood non-mercaptoalbumin/mercaptoalbumin ratio as an oxidative stress marker. METHODS: This study used a crossover design with exercise and control conditions. Under exercise conditions, a bicycle ergometer was used to perform 40 min of transient constant-load exercise at 65% heart rate reserve. Under control conditions, participants rested for 40 min. Blood was collected before, 30 min after, and 90 min after exercise, and at the same time points under control conditions. Samples were analyzed for the homocysteine concentration and non-mercaptoalbumin/mercaptoalbumin ratio. RESULTS: The results revealed that a 65% heart rate reserve and 40 min of acute exercise increased plasma homocysteine concentration and non-mercaptoalbumin ratio. In the intra-condition comparison, the plasma Hcy concentration was significantly increased at Post 30 min (+ 0.83 ± 0.70 µmol/L, P = 0.003) compared with that at Pre in the exercise condition. Furthermore, 90 min after exercise, the blood non-mercaptoalbumin ratio was significantly increased (+ 0.35 ± 0.71%, P = 0.030) compared to Pre. CONCLUSION: These results indicate that the plasma Hcy concentration first increased, and then the non-mercaptoalbumin/mercaptoalbumin ratio increased as the elevated state was maintained. This study revealed that 65% heart rate reserve, 40 min of acute exercise increased plasma Hcy concentration and non-mercaptoalbumin ratio.

Scleral PERK and ATF6 as targets of myopic axial elongation of mouse eyes.

Axial length is the primary determinant of eye size, and it is elongated in myopia. However, the underlying mechanism of the onset and progression of axial elongation remain unclear. Here, we show that endoplasmic reticulum (ER) stress in sclera is an essential regulator of axial elongation in myopia development through activation of both PERK and ATF6 axis followed by scleral collagen remodeling. Mice with lens-induced myopia (LIM) showed ER stress in sclera. Pharmacological interventions for ER stress could induce or inhibit myopia progression. LIM activated all IRE1, PERK and ATF6 axis, and pharmacological inhibition of both PERK and ATF6 suppressed myopia progression, which was confirmed by knocking down above two genes via CRISPR/Cas9 system. LIM dramatically changed the expression of scleral collagen genes responsible for ER stress. Furthermore, collagen fiber thinning and expression of dysregulated collagens in LIM were ameliorated by 4-PBA administration. We demonstrate that scleral ER stress and PERK/ATF6 pathway controls axial elongation during the myopia development in vivo model and 4-PBA eye drop is promising drug for myopia suppression/treatment.

Retinal Degeneration in a Murine Model of Retinal Ischemia by Unilateral Common Carotid Artery Occlusion.

Retinal degeneration is a progressive retinal damage in ocular vascular diseases. There are several reasons for this, such as occlusion of arteries or veins, diabetic retinopathy, or hereditary retinal diseases. To study pathological mechanisms of retinal degeneration, it is required to develop experimentally reproducible and clinically relevant models. In our previous studies, we developed a murine model of retinal hypoperfusion by unilateral common carotid artery occlusion (UCCAO) which mimics the pathophysiology of ocular ischemic syndrome (OIS) in humans, and described broad pathological mechanisms in the retina after UCCAO. However, there still remain missing pieces of the ocular pathologic process by UCCAO. In this study, we examined those unfound mechanisms. UCCAO was performed on adult mice. Ocular dysfunctions, histological deficits, and inflammation were examined after UCCAO, compared with sham-operated mice. Evaluation values were analyzed by electrophysiological, histological, and molecular biological methods. Eyelid drooping was permanently seen after UCCAO. Induction time point of acute reversible cataract under anesthesia was shortened. Retinal/visual dysfunctions were detected 2-4 weeks after UCCAO. Specifically, scotopic b-wave was more affected than a-wave, with the dysfunction of photopic b-wave. Impaired oscillatory potentials and visual evoked potential were constantly observed. Pathological Müller gliosis/inflammation was featured with NeuN-positive cell loss in the ganglion cell layer. Axial length, intraocular pressure, pupillary light reflex, and retinal pigment epithelium/choroidal thickness were not changed by UCCAO. A murine model of retinal ischemia by UCCAO can be useful for studying a series of degenerative process in the ischemic retina.

Salt Restriction Affects the Excretions of Minerals (Na, K, Ca, Mg, P and Zn) in the Second Voided Fasting Early Morning Urine.

The plasma concentrations of mineral (sodium (Na), potassium (K), calcium (Ca), magnesium (Mg), phosphorus (P), and zinc (Zn)) are kept within narrow ranges to maintain homeostasis; hence, it is difficult to use them as indicators of nutritional status. We selected the excretion of these minerals in the second voided fasting early morning urine (EMU) as potential indicators of nutritional status. We previously reported that Na restriction caused a negative balance of Ca and Mg. Therefore, Na restriction can cause changes in EMU-minerals. This study aimed to examine the relationship between dietary Na restriction and urinary mineral excretion. The study lasted for 21 d, including 16 d of balance period and 3 d of recovery period. The participants (11 healthy young women) were divided into the Na restriction group (n=5) (NaCl: 6 g/d) and control group (n=6) (NaCl: 12 g/d). The Na restriction group changed to the control diet (NaCl: 12 g/d) during only the recovery period. The EMU-Na, Ca, Mg, P and Zn in the Na restriction group significantly decreased compared with that of the control group. The EMU-Na, K, Ca, Mg, and Zn in the group with NaCl intake of 6 g/d significantly decreased compared with that of the group with NaCl intake of 12 g/d (in the Na restriction group). We conclude that the decrease in excretion of Na, Ca, Mg and Zn in the EMU can lead to Na restriction. This result can serve as basis when considering EMU as an indicator of mineral status.

Dietary Salt (Sodium Chloride) Requirement and Adverse Effects of Salt Restriction in Humans.

Inevitable sodium loss under sodium restriction must not be construed as evidence for the estimated average requirement (EAR) for sodium (Na) in humans. We conducted human mineral balance studies to determine the EAR for some minerals (Na, K, Ca, Mg, P, Zn, Fe, Cu and Mn). Na concentration in arm sweat was low while those of calcium (Ca) and magnesium (Mg) were high, during relatively heavy bicycle-ergometer exercise under relatively low Na intake (100 mmol/d). This suggests that Na was released from the bone, the sole pool of Na, with Ca and Mg. Additionally, the negative balances of Ca and Mg was observed under a relatively low sodium intake (100 mmol/d) even with the sufficient supply and intake of Ca and Mg into human body. Finally, we found no correlation between the Na intake and the Na balance, while the Na-intake was correlated significantly to the balances of K, Ca and Mg. The Na intake necessary to keep the balances of Ca and Mg positive was calculated to be 68 mg/kg body weight/d. To learn the signs and symptoms of low sodium intake, we compared the results of a metabolic study in which subjects consumed diets with 6 g and 12 g salt/d respectively. The blood pressure decreased only with the 6 g/d group. Fecal moisture contents of the 6 g/d group were lower than for the 12 g/d group, suggesting the fecal Na was strongly reabsorbed with water when the dietary Na was insufficienct. Indiscriminate Na restriction may have adverse effects on health.