Molecular basis of Wnt biogenesis, secretion, and Wnt7-specific signaling.

Wnt proteins are enzymatically lipidated by Porcupine (PORCN) in the ER and bind to Wntless (WLS) for intracellular transport and secretion. Mechanisms governing the transfer of these low-solubility Wnts from the ER to the extracellular space remain unclear. Through structural and functional analyses of Wnt7a, a crucial Wnt involved in central nervous system angiogenesis and blood-brain barrier maintenance, we have elucidated the principles of Wnt biogenesis and Wnt7-specific signaling. The Wnt7a-WLS complex binds to calreticulin (CALR), revealing that CALR functions as a chaperone to facilitate Wnt transfer from PORCN to WLS during Wnt biogenesis. Our structures, functional analyses, and molecular dynamics simulations demonstrate that a phospholipid in the core of Wnt-bound WLS regulates the association and dissociation between Wnt and WLS, suggesting a lipid-mediated Wnt secretion mechanism. Finally, the structure of Wnt7a bound to RECK, a cell-surface Wnt7 co-receptor, reveals how RECKCC4 engages the N-terminal domain of Wnt7a to activate Wnt7-specific signaling.

Mechanisms and inhibition of Porcupine-mediated Wnt acylation.

Wnt signalling is essential for regulation of embryonic development and adult tissue homeostasis1-3, and aberrant Wnt signalling is frequently associated with cancers4. Wnt signalling requires palmitoleoylation on a hairpin 2 motif by the endoplasmic reticulum-resident membrane-bound O-acyltransferase Porcupine5-7 (PORCN). This modification is indispensable for Wnt binding to its receptor Frizzled, which triggers signalling8,9. Here we report four cryo-electron microscopy structures of human PORCN: the complex with the palmitoleoyl-coenzyme A (palmitoleoyl-CoA) substrate; the complex with the PORCN inhibitor LGK974, an anti-cancer drug currently in clinical trials10; the complex with LGK974 and WNT3A hairpin 2 (WNT3Ap); and the complex with a synthetic palmitoleoylated WNT3Ap analogue. The structures reveal that hairpin 2 of WNT3A, which is well conserved in all Wnt ligands, inserts into PORCN from the lumenal side, and the palmitoleoyl-CoA accesses the enzyme from the cytosolic side. The catalytic histidine triggers the transfer of the unsaturated palmitoleoyl group to the target serine on the Wnt hairpin 2, facilitated by the proximity of the two substrates. The inhibitor-bound structure shows that LGK974 occupies the palmitoleoyl-CoA binding site to prevent the reaction. Thus, this work provides a mechanism for Wnt acylation and advances the development of PORCN inhibitors for cancer treatment.

Structure of nevanimibe-bound tetrameric human ACAT1.

Cholesterol is an essential component of mammalian cell membranes, constituting up to 50% of plasma membrane lipids. By contrast, it accounts for only 5% of lipids in the endoplasmic reticulum (ER)1. The ER enzyme sterol O-acyltransferase 1 (also named acyl-coenzyme A:cholesterol acyltransferase, ACAT1) transfers a long-chain fatty acid to cholesterol to form cholesteryl esters that coalesce into cytosolic lipid droplets. Under conditions of cholesterol overload, ACAT1 maintains the low cholesterol concentration of the ER and thereby has an essential role in cholesterol homeostasis2,3. ACAT1 has also been implicated in Alzheimer's disease4, atherosclerosis5 and cancers6. Here we report a cryo-electron microscopy structure of human ACAT1 in complex with nevanimibe7, an inhibitor that is in clinical trials for the treatment of congenital adrenal hyperplasia. The ACAT1 holoenzyme is a tetramer that consists of two homodimers. Each monomer contains nine transmembrane helices (TMs), six of which (TM4-TM9) form a cavity that accommodates nevanimibe and an endogenous acyl-coenzyme A. This cavity also contains a histidine that has previously been identified as essential for catalytic activity8. Our structural data and biochemical analyses provide a physical model to explain the process of cholesterol esterification, as well as details of the interaction between nevanimibe and ACAT1, which may help to accelerate the development of ACAT1 inhibitors to treat related diseases.

Cryo-EM structure of oxysterol-bound human Smoothened coupled to a heterotrimeric Gi.

The oncoprotein Smoothened (SMO), a G-protein-coupled receptor (GPCR) of the Frizzled-class (class-F), transduces the Hedgehog signal from the tumour suppressor Patched-1 (PTCH1) to the glioma-associated-oncogene (GLI) transcription factors, which activates the Hedgehog signalling pathway1,2. It has remained unknown how PTCH1 modulates SMO, how SMO is stimulated to form a complex with heterotrimeric G proteins and whether G-protein coupling contributes to the activation of GLI proteins3. Here we show that 24,25-epoxycholesterol, which we identify as an endogenous ligand of PTCH1, can stimulate Hedgehog signalling in cells and can trigger G-protein signalling via human SMO in vitro. We present a cryo-electron microscopy structure of human SMO bound to 24(S),25-epoxycholesterol and coupled to a heterotrimeric Gi protein. The structure reveals a ligand-binding site for 24(S),25-epoxycholesterol in the 7-transmembrane region, as well as a Gi-coupled activation mechanism of human SMO. Notably, the Gi protein presents a different arrangement from that of class-A GPCR-Gi complexes. Our work provides molecular insights into Hedgehog signal transduction and the activation of a class-F GPCR.

Mechanistic Insights into the Generation and Transduction of Hedgehog Signaling.

Cell differentiation and proliferation require Hedgehog (HH) signaling and aberrant HH signaling causes birth defects or cancers. In this signaling pathway, the N-terminally palmitoylated and C-terminally cholesterylated HH ligand is secreted into the extracellular space with help of the Dispatched-1 (DISP1) and Scube2 proteins. The Patched-1 (PTCH1) protein releases its inhibition of the oncoprotein Smoothened (SMO) after binding the HH ligand, triggering downstream signaling events. In this review, we discuss the recent structural and biochemical studies on four major components of the HH pathway: the HH ligand, DISP1, PTCH1, and SMO. This research provides mechanistic insights into how HH signaling is generated and transduced from the cell surface into the intercellular space and will aid in facilitating the treatment of HH-related diseases.

Mechanisms of Alveolar Type 2 Epithelial Cell Death During Acute Lung Injury.

Diffuse alveolar epithelial cell (AEC) death occurs extensively during acute lung injury (ALI). Due to the limited proliferative capacity of alveolar type 1 epithelial (AT1) cells, the differentiation and regenerative capacity of alveolar type 2 epithelial (AT2) cells are required to restore the barrier function of AECs. However, during lung injury, AT1 cells are particularly susceptible to injury, and ATII cells die in the presence of severe or certain types of injury. This disruption ultimately results in a hindrance to the ability of AT2 cells to proliferate and differentiate into AT1 cells in time to repair the extensively damaged AECs. Therefore, understanding the mechanism of injury death of AT2 cells may be beneficial to reverse the above situation. This article reviews the main death modes of AT2 cells, including apoptosis, necrosis, necroptosis, pyroptosis, autophagic cell death, and ferroptosis. It compares the various forms of death, showing that various cell injury death modes have unique action mechanisms and partially overlapping pathways. Studying the mechanism of AT2 cell death is helpful in screening and analyzing the target pathway of AEC barrier function recovery. It opens up new ideas and strategies for preventing and treating ALI.

Structures of human Patched and its complex with native palmitoylated sonic hedgehog.

Hedgehog (HH) signalling governs embryogenesis and adult tissue homeostasis in mammals and other multicellular organisms1-3. Whereas deficient HH signalling leads to birth defects, unrestrained HH signalling is implicated in human cancers2,4-6. N-terminally palmitoylated HH releases the repression of Patched to the oncoprotein smoothened (SMO); however, the mechanism by which HH recognizes Patched is unclear. Here we report cryo-electron microscopy structures of human patched 1 (PTCH1) alone and in complex with the N-terminal domain of 'native' sonic hedgehog (native SHH-N has both a C-terminal cholesterol and an N-terminal fatty-acid modification), at resolutions of 3.5 Å and 3.8 Å, respectively. The structure of PTCH1 has internal two-fold pseudosymmetry in the transmembrane core, which features a sterol-sensing domain and two homologous extracellular domains, resembling the architecture of Niemann-Pick C1 (NPC1) protein7. The palmitoylated N terminus of SHH-N inserts into a cavity between the extracellular domains of PTCH1 and dominates the PTCH1-SHH-N interface, which is distinct from that reported for SHH-N co-receptors8. Our biochemical assays show that SHH-N may use another interface, one that is required for its co-receptor binding, to recruit PTCH1 in the absence of a covalently attached palmitate. Our work provides atomic insights into the recognition of the N-terminal domain of HH (HH-N) by PTCH1, offers a structural basis for cooperative binding of HH-N to various receptors and serves as a molecular framework for HH signalling and its malfunction in disease.

Molecular basis of cholesterol efflux via ABCG subfamily transporters.

The ABCG1 homodimer (G1) and ABCG5-ABCG8 heterodimer (G5G8), two members of the adenosine triphosphate (ATP)-binding cassette (ABC) transporter G family, are required for maintenance of cellular cholesterol levels. G5G8 mediates secretion of neutral sterols into bile and the gut lumen, whereas G1 transports cholesterol from macrophages to high-density lipoproteins (HDLs). The mechanisms used by G5G8 and G1 to recognize and export sterols remain unclear. Here, we report cryoelectron microscopy (cryo-EM) structures of human G5G8 in sterol-bound and human G1 in cholesterol- and ATP-bound states. Both transporters have a sterol-binding site that is accessible from the cytosolic leaflet. A second site is present midway through the transmembrane domains of G5G8. The Walker A motif of G8 adopts a unique conformation that accounts for the marked asymmetry in ATPase activities between the two nucleotide-binding sites of G5G8. These structures, along with functional validation studies, provide a mechanistic framework for understanding cholesterol efflux via ABC transporters.

Regulating the Singlet and Triplet Emission of Sb3+ Ions to Achieve Single-Component White-Light Emitter with Record High Color-Rendering Index and Stability.

The rapid development of solid-state lighting technology has attracted much attention for searching efficient and stable luminescent materials, especially the single-component white-light emitter. Here, we adopt a facile ion-doping technology to synthesize vacancy-ordered double perovskite Cs2ZrCl6:Sb. The introduction of Sb3+ ions with a 5s2 active lone pair into Cs2ZrCl6 host stimulates the singlet (blue) and triplet (orange) states emission of Sb3+ ions, and their relative emission intensity can be tuned through the energy transfer from singlet to triplet states. Benefiting from the dual-band emission as a pair of perfect complementary colors, the optimum Cs2ZrCl6:1.5%Sb exhibits a high-quality white emission with a color-rendering index of 96. By employing Cs2ZrCl6:1.5%Sb as the down-conversion phosphor, stable single-component white light-emitting diodes with a record half-lifetime of 2003 h were further fabricated. This study puts forward an effective ion-doping strategy to design single-component white-light emitter, making practical applications of them in lighting technologies a real possibility.

Sterols in an intramolecular channel of Smoothened mediate Hedgehog signaling.

Smoothened (SMO), a class Frizzled G protein-coupled receptor (class F GPCR), transduces the Hedgehog signal across the cell membrane. Sterols can bind to its extracellular cysteine-rich domain (CRD) and to several sites in the seven transmembrane helices (7-TMs) of SMO. However, the mechanism by which sterols regulate SMO via multiple sites is unknown. Here we determined the structures of SMO-Gi complexes bound to the synthetic SMO agonist (SAG) and to 24(S),25-epoxycholesterol (24(S),25-EC). A novel sterol-binding site in the extracellular extension of TM6 was revealed to connect other sites in 7-TMs and CRD, forming an intramolecular sterol channel from the middle side of 7-TMs to CRD. Additional structures of two gain-of-function variants, SMOD384R and SMOG111C/I496C, showed that blocking the channel at its midpoints allows sterols to occupy the binding sites in 7-TMs, thereby activating SMO. These data indicate that sterol transport through the core of SMO is a major regulator of SMO-mediated signaling.

[A rare case of dicentric ring chromosome and derivative ring chromosome Chimera].

OBJECTIVE: Utilize high-resolution chromosome analysis and microarray detection to determine the genetic etiology of infertility of a 32-year old female patient. METHODS: The peripheral blood of the patient was cultured for high-resolution chromosome G and C banding karyotype analysis, and then 750K SNP-Array chip detection was performed. RESULTS: Karyotype analysis results showed that the patient's karyotype was 45,XX,-13 [7]/46,XX,r(13) (p13q34) [185]/46,XX,dic r(13;13)(p13q34;p13q34) [14]/ 47,XX,+der(13;13;13;13) (p13q34;p13q34;p13q34; p13q34), dic r(13;13) [1]/ 46,XX [3]. The microarray results showed that the patient had a 3.3 Mb deletion in the 13q34 segment of chromosome 13, which may be related to infertility. CONCLUSION: Infertility of the patient reported in this article may be related to the deletion of chromosome segment (13q34-qter).

Urban-Rural Differences in Bone Mineral Density and its Association with Reproductive and Menstrual Factors Among Older Women.

PURPOSE: This study aimed to compare the bone mineral density (BMD) of older women living in rural and urban areas, and evaluate the potential factors affecting the risk of osteoporosis. METHODS: We recruited 574 women aged 65 years or older from rural areas and 496 from urban areas in Shanghai, China. The BMD values of the lumbar vertebrae and total left hip were measured by a dual energy X-ray absorptiometry densitometer. We also recorded information about education level, family income, medications, reproductive and menstrual history, diet, smoking, and alcohol consumption. RESULTS: Women in urban areas had significantly higher BMD in their lumbar spine, and there was a dramatic increase in the proportion of women with osteoporosis in rural areas. The age at menarche was significantly higher among women living in rural areas, and there were more years from menarche to menopause among urban women. Rural women had significantly higher numbers of both pregnancies and parity, and a significantly lower age at first parity. In multiple linear regression analyses, years from menarche to menopause was independently related to high lumbar spine BMD, while age at menarche and parity was independently related to low lumbar spine BMD. CONCLUSION: More older women in rural areas had osteoporosis. Later menarche, less years from menarche to menopause and higher parity might partially contribute to decreased BMD among women in rural areas. More attention should be paid to women in rural areas to prevent bone loss and further bone and health impairment.

Structural basis of rifampin inactivation by rifampin phosphotransferase.

Rifampin (RIF) is a first-line drug used for the treatment of tuberculosis and other bacterial infections. Various RIF resistance mechanisms have been reported, and recently an RIF-inactivation enzyme, RIF phosphotransferase (RPH), was reported to phosphorylate RIF at its C21 hydroxyl at the cost of ATP. However, the underlying molecular mechanism remained unknown. Here, we solve the structures of RPH from Listeria monocytogenes (LmRPH) in different conformations. LmRPH comprises three domains: an ATP-binding domain (AD), an RIF-binding domain (RD), and a catalytic His-containing domain (HD). Structural analyses reveal that the C-terminal HD can swing between the AD and RD, like a toggle switch, to transfer phosphate. In addition to its catalytic role, the HD can bind to the AD and induce conformational changes that stabilize ATP binding, and the binding of the HD to the RD is required for the formation of the RIF-binding pocket. A line of hydrophobic residues forms the RIF-binding pocket and interacts with the 1-amino, 2-naphthol, 4-sulfonic acid and naphthol moieties of RIF. The R group of RIF points toward the outside of the pocket, explaining the low substrate selectivity of RPH. Four residues near the C21 hydroxyl of RIF, His825, Arg666, Lys670, and Gln337, were found to play essential roles in the phosphorylation of RIF; among these the His825 residue may function as the phosphate acceptor and donor. Our study reveals the molecular mechanism of RIF phosphorylation catalyzed by RPH and will guide the development of a new generation of rifamycins.

Nerve Growth Factor Changes and Corneal Nerve Repair after Keratoplasty.

SIGNIFICANCE: Measured tear concentration of nerve growth factor is correlated with postoperative corneal reinnervation among patients who undergo keratoplasty. This may be a future therapeutic target for post-keratoplasty corneal nerve regeneration. PURPOSE: To determine the relationship between changes in the content of nerve growth factor (NGF) in tear fluid and corneal subepithelial nerve regeneration in patients after keratoplasty. METHODS: In this retrospective study, 30 eyes of 28 patients (15 males, 13 females; mean age 42.8 [range 16-73] years) who underwent primary keratoplasty for the first time were recruited through the clinics of the Department of Ophthalmology, Jilin University affiliated First Hospital, between May and December 2015. All patients underwent a complete ophthalmic examination preoperatively. Tear fluid samples were collected to detect the content of NFG at different time points in the follow-up period (day 1 preoperatively and days 1, 7, 30, and 90 postoperatively) and analyzed correlations between NFG content and age, infective factors, and variables of the surgical procedure as well as with subepithelial nerve repair at 30 and 90 days postoperatively. RESULTS: The NFG content in tear fluid on day 1 postoperatively was lower than that on the day preceding surgery; however, it was higher than the preoperative value on postoperative days 7, 30, and 90 (F = 5.046, P < 0.05). Further, the NFG content of tear fluid at 30 days postoperatively correlated with the surgical procedure (coefficient = -2.775, P = 0.010); however, no significant correlation was found on postoperative day 1 (coefficient = -1.315, P > 0.05). At all study time points, the NFG content of tear fluid had no correlation with infective factors or age (P > 0.05). Postoperatively, at day 30, small nerve buds were observed in the periphery of the corneal graft in 13 eyes (43.3% of cases) but not in 17 eyes (56.7% of cases), which showed a significant correlation with the NFG content of tear fluid (coefficient = -3.370, P = 0.010). By postoperative day 90, small nerve buds were observed in the periphery of the corneal graft in 24 eyes (80.0% of cases) and showed a significant correlation with the NFG content of tear fluid (coefficient = -2.750, P = 0.006). CONCLUSIONS: The NFG content in tear fluid increases with the increasing ratio of small nerve buds indicating corneal nerve regeneration. NFG promotes subepithelial nerve regeneration in patients after keratoplasty.

Impact of tumor architecture on disease recurrence and cancer-specific mortality of upper tract urothelial carcinoma treated with radical nephroureterectomy.

Upper tract urinary carcinoma (UTUC) is a relatively uncommon but aggressive disease. Recent publications have assessed the prognostic significance of tumor architecture in UTUC, but there is still controversy regarding the significance and importance of tumor architecture on disease recurrence. We retrospectively reviewed the medical records of 101 patients with clinical UTUC who had undergone surgery. Univariate and multivariate analyses were conducted to identify factors associated with disease recurrence and cancer-specific mortality. As our single center study and the limited sample size may influence the clinical significance, we further quantitatively combined the results with those of existing published literature through a meta-analysis compiled from searching several databases. At a median follow-up of 41.3 months, 25 patients experienced disease recurrence. Spearman's correlation analysis showed that tumor architecture was found to be positively correlated with the tumor location and the histological grade. Kaplan-Meier curves showed that patients with sessile tumor architecture had significantly poor recurrence free survival (RFS) and cancer specific survival (CSS). Furthermore, multivariate analysis suggested that tumor architecture was independent prognostic factors for RFS (Hazard ratio, HR = 2.648) and CSS (HR = 2.072) in UTUC patients. A meta-analysis of investigating tumor architecture and its effects on UTUC prognosis was conducted. After searching PubMed, Medline, Embase, Cochrane Library and Scopus databases, 17 articles met the eligibility criteria for this analysis. The eligible studies included a total of 14,368 patients and combined results showed that sessile tumor architecture was associated with both disease recurrence with a pooled HR estimate of 1.454 and cancer-specific mortality with a pooled HR estimate of 1.416. Tumor architecture is an independent predictor for disease recurrence after radical nephroureterectomy for UTUC. Therefore, closer surveillance is necessary, especially in patients with sessile tumor architecture.

Structural basis of intramitochondrial phosphatidic acid transport mediated by Ups1-Mdm35 complex.

Ups1 forms a complex with Mdm35 and is critical for the transport of phosphatidic acid (PA) from the mitochondrial outer membrane to the inner membrane. We report the crystal structure of the Ups1-Mdm35-PA complex and the functional characterization of Ups1-Mdm35 in PA binding and transfer. Ups1 features a barrel-like structure consisting of an antiparallel ß-sheet and three α-helices. Mdm35 adopts a three-helical clamp-like structure to wrap around Ups1 to form a stable complex. The ß-sheet and α-helices of Ups1 form a long tunnel-like pocket to accommodate the substrate PA, and a short helix α2 acts as a lid to cover the pocket. The hydrophobic residues lining the pocket and helix α2 are critical for PA binding and transfer. In addition, a hydrophilic patch on the surface of Ups1 near the PA phosphate-binding site also plays an important role in the function of Ups1-Mdm35. Our study reveals the molecular basis of the function of Ups1-Mdm35 and sheds new light on the mechanism of intramitochondrial phospholipid transport by the MSF1/PRELI family proteins.

Adaptive optics retinal imaging with automatic detection of the pupil and its boundary in real time using Shack-Hartmann images.

Retinal imaging with an adaptive optics (AO) system usually requires that the eye be centered and stable relative to the exit pupil of the system. Aberrations are then typically corrected inside a fixed circular pupil. This approach can be restrictive when imaging some subjects, since the pupil may not be round and maintaining a stable head position can be difficult. In this paper, we present an automatic algorithm that relaxes these constraints. An image quality metric is computed for each spot of the Shack-Hartmann image to detect the pupil and its boundary, and the control algorithm is applied only to regions within the subject's pupil. Images on a model eye as well as for five subjects were obtained to show that a system exit pupil larger than the subject's eye pupil could be used for AO retinal imaging without a reduction in image quality. This algorithm automates the task of selecting pupil size. It also may relax constraints on centering the subject's pupil and on the shape of the pupil.

In vivo confocal microscopy of pre-Descemet corneal dystrophy associated with X-linked ichthyosis: a case report.

BACKGROUND: Pre-Descemet corneal dystrophy (PDCD) is characterized by the presence of numerous, tiny, polymorphic opacities immediately anterior to Descemet membrane, which is a rare form of corneal stromal dystrophy and hard to be diagnosed. In vivo confocal microscopy (IVCM) is a useful tool to examine the minimal lesions of the cornea at the cellular level. In this article, we report a rare case of PDCD associated with X-linked ichthyosis and evaluate IVCM findings. CASE PRESENTATION: We present a 34-year-old male Chinese patient with PDCD associated with X-linked ichthyosis. Slit-lamp biomicroscopy showed the presence of tiny and pleomorphic opacities in the posterior stroma immediately anterior to Descemet membrane bilaterally. IVCM revealed regular distributed hyperreflective particles inside the enlarged and activated keratocytes in the posterior stroma. Hyperreflective particles were also observed dispersedly outside the keratocytes in the anterior stroma. Dermatological examination revealed that the skin over the patient's entire body was dry and coarse, with thickening and scaling of the skin in the extensor side of the extremities. PCR results demonstrated that all ten exons and part flanking sequences of STS gene failed to produce any amplicons in the patient. CONCLUSIONS: IVCM is useful for analyzing the living corneal structural changes in rare corneal dystrophies. We first reported the IVCM characteristics of PDCD associated with X-linked ichthyosis, which was caused by a deletion of the steroid sulfatase (STS) gene, confirmed by gene analysis.

Investigation of evanescent coupling between tapered fiber and a multimode slab waveguide.

A tapered fiber-slab waveguide coupler (TFSC) is proposed in this paper. Both the numerical analysis based on the beam propagation method and experiments are used for investigating the dependencies of TFSC transmission features on their geometric parameters. From the simulations and experimental results, the rules for fabricating a TFSC with low transmission loss and sharp resonant spectra by optimizing the configuration parameters are presented. The conclusions derived from our work may provide helpful references for optimally designing and fabricating TFSC-based devices, such as sensors, wavelength filters, and intensity modulators.

A clinical planning module for adaptive optics SLO imaging.

PURPOSE: To develop a clinical planning module (CPM) to improve the efficiency of imaging subjects with a steerable wide-field adaptive optics scanning laser ophthalmoscope (AOSLO) and to evaluate the performance of this module by imaging the retina in healthy and diseased eyes. METHODS: We developed a software-based CPM with two submodules: a navigation module and a montage acquisition module. The navigation module guides the AOSLO to image identified retinal regions from a clinical imaging platform using a matrix-based mapping between the two. The montage acquisition module systematically moves the AOSLO steering mirrors across the retina in predefined patterns. The CPM was calibrated using a model eye and tested on five normal subjects and one patient with a retinal nerve fiber layer defect. RESULTS: Within the central ±7° from the fixation target, the CPM can direct the AOSLO beam to the desired regions with localization errors of <0.3°. The navigation error increases with eccentricity, and larger errors (up to 0.8°) were evident for regions beyond 7°. The repeatability of CPM navigation was tested on the same locations from two subjects. The localization errors between trials on different days did not differ significantly (p > 0.05). The region with a size of approximately 13° × 10° can be imaged in about 30 min. An approximately 12° × 4.5° montage of the diseased region from a patient was imaged in 18 min. CONCLUSIONS: We have implemented a clinical planning module to accurately guide the AOSLO imaging beam to desired locations and to quickly acquire high-resolution AOSLO montages. The approach is not only friendly for patients and clinicians but also convenient to relate the imaging data between different imaging platforms.