Intravenous indocyanine green dye is insufficient for robust immune cell labelling in the human retina

It is not currently possible to reliably visualise and track immune cells in the human central nervous system or eye. Previous work demonstrated that indocyanine green (ICG) dye could label immune cells and be imaged after a delay during disease in the mouse retina. We report a pilot study investigating if ICG can similarly label immune cells within the human retina. Twelve adult participants receiving ICG angiography as part of routine standard of care were recruited. Baseline retinal images were obtained prior to ICG administration then repeated over a period ranging from 2 hours to 9 days. Matched peripheral blood samples were obtained to examine systemic immune cell labelling and activation from ICG by flow cytometry with human macrophage cultures as positive controls. Differences between the delayed near infrared ICG imaging and 488 nm autofluorescence was observed across pathologies, likely arising from the retinal pigment epithelium (RPE). Only one subject demonstrated ICG signal on peripheral blood myeloid cells and only three distinct cell-sized signals appeared over time within the retina of three participants. No significant increase in immune cell activation markers were detected after ICG administration. ICG accumulated in the endosomes of macrophage cultures and was detectable above a minimum concentration, suggesting cell labelling is possible. ICG can label RPE and may be used as an additional biomarker for RPE health across a range of retinal disorders. Standard clinical doses of intravenous ICG do not lead to robust immune cell labelling in human blood or retina and further optimisation in dose and route are required

Intravenous indocyanine green dye is insufficient for robust immune cell labelling in the human retina

It is not currently possible to reliably visualise and track immune cells in the human central nervous system or eye. Previous work demonstrated that indocyanine green (ICG) dye could label immune cells and be imaged after a delay during disease in the mouse retina. We report a pilot study investigating if ICG can similarly label immune cells within the human retina. Twelve adult participants receiving ICG angiography as part of routine standard of care were recruited. Baseline retinal images were obtained prior to ICG administration then repeated over a period ranging from 2 hours to 9 days. Matched peripheral blood samples were obtained to examine systemic immune cell labelling and activation from ICG by flow cytometry with human macrophage cultures as positive controls. Differences between the delayed near infrared ICG imaging and 488 nm autofluorescence was observed across pathologies, likely arising from the retinal pigment epithelium (RPE). Only one subject demonstrated ICG signal on peripheral blood myeloid cells and only three distinct cell-sized signals appeared over time within the retina of three participants. No significant increase in immune cell activation markers were detected after ICG administration. ICG accumulated in the endosomes of macrophage cultures and was detectable above a minimum concentration, suggesting cell labelling is possible. ICG can label RPE and may be used as an additional biomarker for RPE health across a range of retinal disorders. Standard clinical doses of intravenous ICG do not lead to robust immune cell labelling in human blood or retina and further optimisation in dose and route are required

The actions of prostaglandins on the urinary bladder

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Prostaglandin E(2) induces spontaneous rhythmic activity in mouse urinary bladder independently of efferent nerves.

Background and purpose:  The acute effects of PGE(2) on bladder smooth muscle and nerves were examined to determine the origin of PGE(2) -induced spontaneous rhythmic contractions. Experimental Approach:  Contraction studies, confocal Ca(2+) imaging and electrophysiology in strips of mouse urinary bladder. Key Results:  Detrusor smooth muscle strips generated a PGE(2)  (50 µM)-induced increase in tone and phasic contractions, a characteristic of detrusor overactivity. Confocal Ca(2+) imaging showed a PGE(2) -induced increase in the frequency of whole cell Ca(2+) transients (WCTs) (72 ± 5%) and intracellular recordings showed an increased frequency of spontaneous depolarizations, from 0.31 s(-1) to 0.90 s(-1) . Non-selective inhibition of EP receptors using SC-51322 and AH-6809 (10 µM), or the L-type Ca(2+) channel blocker nifedipine (1 µM), prevented phasic contractions and WCTs, and reduced the tone (by 45 ± 7% and 59 ± 6% respectively). Blocking P2X(1) receptors with NF449 (10 µM) caused a small but significant reduction in the frequency of PGE(2) -induced phasic contractions (24 ± 9%) and WCTs (28 ± 17%) but had no significant effect on spontaneous depolarizations or tone. Inhibiting muscarinic receptors with cyclopentolate (1 µM) had no significant effect on these measures. Spontaneous WCTs became synchronous in PGE(2) , implying enhanced functional coupling between neighbouring cells. However, the electrical input resistance was unchanged (median control 181 MΩ; PGE(2) median 209 MΩ). Conclusions and Implications:  It is argued that depolarization alone is sufficient to explain a functional increase in intercellular coupling. The action of PGE(2 ) to increase detrusor spontaneous rhythmic activity does not require parasympathetic nerves

Prostaglandin E2 induces spontaneous rhythmic activity in mouse urinary bladder independently of efferent nerves

BACKGROUND AND PURPOSE: The acute effects of PGE(2) on bladder smooth muscle and nerves were examined to determine the origin of PGE(2)-induced spontaneous rhythmic contractions. EXPERIMENTAL APPROACH: Contraction studies, confocal Ca(2+) imaging and electrophysiological recordings in strips of mouse urinary bladder were used to differentiate the effects of PGE(2) on bladder smooth muscle and efferent nerves. KEY RESULTS: PGE(2) (50 µM) increased the tone and caused phasic contractions of detrusor smooth muscle strips. Confocal Ca(2+) imaging showed that PGE(2) increased the frequency of whole-cell Ca(2+) transients (WCTs) (72 ± 5%) and intracellular recordings showed it increased the frequency of spontaneous depolarizations, from 0.31·s(−1) to 0.90·s(−1). Non-selective inhibition of EP receptors using SC-51322 and AH-6809 (10 µM), or the L-type Ca(2+) channel blocker nifedipine (1 µM), prevented these phasic contractions and WCTs, and reduced the tone (by 45 ± 7% and 59 ± 6%, respectively). Blocking P2X1 receptors with NF449 (10 µM) caused a small but significant reduction in the frequency of PGE(2)-induced phasic contractions (24 ± 9%) and WCTs (28 ± 17%) but had no significant effect on spontaneous depolarizations or tone. Inhibiting muscarinic receptors with cyclopentolate (1 µM) had no significant effect on these measures. Spontaneous WCTs became synchronous in PGE(2), implying enhanced functional coupling between neighbouring cells. However, the electrical input resistance was unchanged. CONCLUSIONS AND IMPLICATIONS: It was concluded that depolarization alone is sufficient to explain a functional increase in intercellular coupling and the ability of PGE(2) to increase detrusor spontaneous rhythmic activity does not require parasympathetic nerves