Secondary follicles enable efficient germline mtDNA base editing at hard-to-edit site.

Efficient germline mtDNA editing is required to construct disease-related animal models and future gene therapy. Recently, the DddA-derived cytosine base editors (DdCBEs) have made mitochondrial genome (mtDNA) precise editing possible. However, there still exist challenges for editing some mtDNA sites in germline via zygote injection, probably due to the suspended mtDNA replication during preimplantation development. Here, we introduce a germline mtDNA base editing strategy: injecting DdCBEs into oocytes of secondary follicles, at which stage mtDNA replicates actively. With this method, we successfully observed efficient G-to-A conversion at a hard-to-edit site and also obtained live animal models. In addition, for those editable sites, this strategy can greatly improve the base editing efficiency up to 3-fold, which is more than that in zygotes. More important, editing in secondary follicles did not increase more the risk of off-target effects than that in zygotes. This strategy provides an option to efficiently manipulate mtDNA sites in germline, especially for hard-to-edit sites.

scRNA-Seq: First Atlas and Cellular Landscape of Lacrimal Sac: Implications in Primary Acquired Nasolacrimal Duct Obstruction Pathogenesis.

Purpose: The aim of this study was to describe the transcriptional changes of individual cellular components in the lacrimal sac in patients with primary acquired nasolacrimal duct obstruction (PANDO) and attempt to construct the first lacrimal sac cellular atlas to elucidate the potential mechanisms that may drive the disease pathogenesis. Methods: Lacrimal sac samples were obtained intra-operatively during the endoscopic dacryocystorhinostomy (EnDCR) procedure from five patients. Single-cell RNA sequencing was performed to analyze each individual cell population including epithelial and immune cells during the early inflammatory and late inflammatory phases of the disease. Results: Eleven cell types were identified among 25,791 cells. T cells and B cells were the cell populations with the greatest variation in cell numbers between the two phases and were involved in immune response and epithelium migration-related pathways. The present study showed that epithelial cells highly expressed the genes of senescence-associated secretory phenotype (SASP) and were involved in influencing the inflammation, neutrophil chemotaxis, and migration during the late inflammatory stage. Enhanced activity of CXCLs-CXCRs between the epithelial cells and neutrophils was noted by the cell-cell communication analysis and is suspected to play a role in inflammation by recruiting more neutrophils. Conclusions: The study presents a comprehensive single-cell landscape of the lacrimal sac cells in different phases of PANDO. The contribution of T cells, B cells, and epithelial cells to the inflammatory response, and construction of the intercellular signaling networks between the cells within the lacrimal sac has further enhanced the present understanding of the PANDO pathogenesis.

Primary Lacrimal Sac Tumors with Extension into Vicinity: Outcomes with Endoscopy-Assisted Modified Weber-Ferguson's Approach.

PURPOSE: To evaluate the outcomes of endoscopy-assisted modified Weber-Ferguson's approach in the management of primary lacrimal sac tumors with extension into the neighboring tissues. METHODS: A retrospective interventional study was performed on all patients with lacrimal sac tumors treated with the endoscopy-assisted modified Weber-Ferguson approach between January 2010 and June 2022 at the Shanghai Ninth People's Hospital, China. Data assessed include demographics, clinical presentations, imaging features, surgical techniques, histopathology, adjuvant modalities of management, complications, and outcomes. RESULTS: A total of 13 patients were included in the analysis. Epiphora and palpable mass lesion were the presenting complaint in 84.6% (11/13) of the patients. Nearly half of the patients (46.1%, 6/13) were misdiagnosed as lacrimal duct obstruction. All the lacrimal sac tumors in the present series showed uneven enhancement on T1-weighted MRI imaging. Postoperatively, 84.6% (11/13) patients recovered well with excellent esthetics and were disease-free after a mean follow-up of 58.6 months. Two patients who underwent additional exenteration developed recurrence and succumbed (at 41 and 96 months follow up) while they were on palliative chemoradiation. CONCLUSION: The endoscopic-assisted modified Weber-Fergusson surgical approach is effective in providing better visibility and accessibility to lacrimal sac tumors with extension into neighboring tissue.

Evaluation of primary acquired nasolacrimal duct obstruction: Comparison of CT-DCG and dacryoendoscopy in accurately localizing the lacrimal drainage obstructions.

OBJECTIVE: To correlate and evaluate the power and limitations of CT-DCG in determining the level and type of lacrimal duct obstruction in comparison to dacryoendoscopy in patients clinically suspected to be having partial or complete primary acquired nasolacrimal duct obstruction (PANDO). METHODS: A retrospective chart review was performed on 1232 lacrimal drainage systems of 957 patients who suffered from primary acquired nasolacrimal duct obstruction (PANDO) at Shanghai Ninth People's Hospital. Patients were examined with CT-DCG and correlated with dacryoendoscopy and the findings of clinical examination. RESULTS: Of the studied patients, 173 were men and 784 were women with an age range of 18-93 years. Of the 1232 lacrimal pathways, good CT-DCG images could be obtained in 980 cases and dacryoendoscopy in 957 cases. Of these complete obstructions were noted in 81% (794/980), and partial obstructions were identified in 19% (186/980) with CT-DCG. CT-DCG and dacryoendoscopy showed 68.4% agreement for the type of the obstruction and 63% for the level of the obstruction. The majority of the obstructions occurred at the sac-duct junction (62.5%) followed by the upper half of the nasolacrimal duct (27.5%). There was a significant difference in the correlation of the obstruction type with age group and with the duration of symptoms. As the duration of symptoms increased, the proportion of complete lacrimal duct obstructions as shown on CT-DCG images increased and the proportion of incomplete obstruction decreased (p = 0.015). CONCLUSIONS: The junction of lacrimal sac and nasolacrimal duct was the most common obstruction site. Age and the duration of symptoms influenced the type of obstruction noted. The degree and level of agreement between the investigations was moderate. A combination of CT-DCG and Dacryoendoscopy could together identify the location more accurately.

Prognostic factors of traumatic optic neuropathy based on multimodal analysis-Especially the influence of postoperative dressing change and optic nerve blood supply on prognosis.

Objective: To investigate the critical prognostic factors of patients with traumatic optic neuropathy (TON) treated with endoscopic transnasal optic canal decompression (ETOCD) and to perform multimodal analysis based on imaging examinations of optical coherence tomography angiography (OCTA) and CT scan. Subsequently, a new prediction model was established. Methods: The clinical data of 76 patients with TON who underwent decompression surgery with the endoscope-navigation system in the Department of Ophthalmology, Shanghai Ninth People's Hospital from January 2018 to December 2021 were retrospectively analyzed. The clinical data included demographic characteristics, reasons for injury, interval between injury and surgery, multimode imaging information of CT scan and OCTA, including orbital fracture, optical canal fractures, vessel density of optic disc and macula, and the times of postoperative dressing change. Binary logistic regression was used to establish a model for best corrected visual acuity (BCVA) after treatment as a predictor of TON outcome. Results: Postoperative BCVA improved in 60.5% (46/76) patients and did not improve in 39.5% (30/76) patients. The times of postoperative dressing change had a significant impact on the prognosis. Other factors affecting the prognosis were microvessel density of the central optic disc, the cause of injury, and the microvessel density above the macula. The area under the raw current curves of the predictive model was 0.7596. Conclusions: The times of dressing changes after the operation, i.e., continuous treatment, is the key factor affecting prognosis. The microvessel density in the center of the optic disc and superior macula, quantitatively analyzed by OCTA, is the prognostic factor of TON and may be used as a prognostic marker of TON.

Long-term outcomes of a new anatomy-based method for finding the medial cut end during late canalicular repair.

BACKGROUND: Locating the medial cut end during late repair of canalicular lacerations can be challenging. OBJECTIVE: The aim of this study was to evaluate the effectiveness and long-term outcomes of a new anatomy-based method for solving the problem of locating the medial cut end. METHODS: This retrospective interventional study included 85 eyes of 85 consecutive adult patients with unilateral inferior canalicular lacerations who underwent late primary (≥2 days after injury) or secondary (≥6 months after initial treatment) surgery. Before surgery, the lacerations were classified as lateral, central, or medial according to the 'distance from the punctum to the distal end' of the lacerated inferior canaliculus. The time spent to locate the proximal lacerated end (TSL) was recorded. All patients were followed up for ≥1 year to evaluate the lacrimal passage patency and the distance between the superior and inferior punctum (DBSIP, to assess cosmesis). RESULTS: There were 16 (18.82%) lateral-type, 55 (64.71%) central-type, and 14 (16.47%) medial-type canalicular lacerations. The TSL was 3.48 ± 1.05 (range 0.9-6.8) min for all patients and differed significantly among the three types of canalicular lacerations (P < 0.001). Lacrimal irrigation showed patent lacrimal passages in 69 patients (81.18%) at 3 months and a further 4 patients (4.71%) at 6 months, residual stenosis without obstruction in 5 patients (5.88%), and obstruction in 7 patients (8.24%). The postoperative DBSIP on the affected side was shorter than the preoperative DBSIP (2.66 ± 0.66 vs. 3.09 ± 1.72 mm, P = 0.006) and comparable to that on the unaffected side (2.78 ± 0.40 mm). CONCLUSION: Our new anatomy-based method is efficient and achieves good long-term outcomes for all types of late canalicular repair.

Targeted OUM1/PTPRZ1 silencing and synergetic CDT/enhanced chemical therapy toward uveal melanoma based on a dual-modal imaging-guided manganese metal-organic framework nanoparticles.

Metastasis and chemical resistance are the most serious problems in the treatment of highly aggressive uveal melanoma (UM). The newly identified lncRNA OUM1 is overexpressed in UM, functions as a catalyst and regulates protein tyrosine phosphatase (PTP) activity by binding to PTP receptor type Z1 (PTPRZ1), which plays an important role in cell proliferation, metastasis and chemotherapy resistance in the UM microenvironment. Hence, siRNAs that selectively knocking down the lncRNA OUM1 (siOUM1) and its target gene PTPRZ1 (siPTPRZ1) were designed to inhibit the OUM1/PTPRZ1 pathway to reduce PTP activity, and this reduction in activity interrupts protein tyrosine phosphorylation, suppresses UM proliferation and metastasis and improves cisplatin sensitivity in UM cells. Then, to overcome the limitations of the difficulty of drug administration and traditional therapeutics, the indocyanine green (ICG)-labeled manganese metal-organic framework (MOF) nanoparticles (NPs) were fabricated and linked with arginine-glycine-aspartate (RGD) peptide to carry siOUM1/siPTPRZ1 and cisplatin to achieve targeted siRNA interference-mediated therapy, enhanced cisplatin therapy and chemodynamic therapy. This NP system also has a dual-modal imaging ability because ICG is a near-infrared region fluorescent dye and manganese has the potential to be used in magnetic resonance imaging. This study verifies the significance of the newly discovered lncRNA OUM1 as a new therapeutic target for aggressive UM and provides a drug delivery NP system for precise treatment of UM accompanied with a dual-modal imaging ability.

Differently Expressed Circular RNAs in Lacrimal Sacs From Patients With Chronic Dacryocystitis.

This study was designed to identify differently expressed circular RNAs (circRNAs) and investigate their potential roles in lacrimal sacs from patients with chronic dacryocystitis. The lacrimal sac samples of three chronic dacryocystitis patients and three control subjects were collected for RNA sequencing after ribosomal RNA was depleted. Differently expressed circRNAs and messenger RNAs (mRNAs) were used for co-expression analysis. CircRNA-microRNA (miRNA)-mRNA interaction network were also established by miRanda software. Meanwhile, pathway and functional enrichment analysis were conducted for the down- and up-regulated mRNAs in the circRNA-mRNA co-expression network. The expression levels of circRNAs and mRNAs in chronic dacryocystitis and control samples were validated by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). In all the 3,909 circRNAs predicted through RNA sequencing, 25 circRNAs (20 up-regulated and 5 down-regulated) expressed differently in chronic dacryocystitis samples. Besides, there identified 1,486 differentially expressed mRNAs. Of these differently expressed circRNAs and mRNAs, eight were validated by qRT-PCR, including MYH2, DSP, CD27, CCL5, FN1, has_circ_0004792, has_circ_0001062, and has_circ_0115476. Gene Ontology (GO) analysis indicated that the majority of altered mRNAs in this co-expression network were involved in immune system processes and meanwhile Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that these altered expressed mRNAs were also amplified in bacterial invasion of epithelial cells, both of which were thought to be involved in the pathogenesis of chronic dacryocystitis. In the circRNA-miRNA-mRNA interaction network, six circRNAs were found to be related to Th1 and Th2 cell differentiation, which was closely associated with the development of chronic dacryocystitis. This study identified statistically significant differences between circRNAs and mRNAs of lacrimal sac samples of chronic dacryocystitis patients and control individuals and provides novel insight into the regulatory mechanism of circRNAs, miRNAs, and mRNAs in the pathogenesis of chronic dacryocystitis.

Noninvasive Machine Learning Screening Model for Dacryocystitis Based on Ocular Surface Indicators.

BACKGROUND: Dacryocystitis is an orbital disease that can be easily misdiagnosed. The most common diagnostic tools for dacryocystitis are computed tomography, lacrimal duct angiography, and lacrimal tract irrigation. Yet, those are invasive methods, which are not conducive to extensive screening. OBJECTIVE: To explore the significance of ocular surface indicators and demographic data in the screening of dacryocystitis. MATERIALS AND METHODS: Data were prospectively collected from 56 patients with dacryocystitis (56 eyes) and 56 healthy individuals. Collected indicators included demographic information (gender, age), ocular surface data of tear meniscus height, objective scatter index (OSI), and clinical diagnosis. The model features were screened out by machine learning to establish a dacryocystitis screening model. RESULTS: Tear meniscus height, OSI_maximum Lyapunov exponent, basic OSI, median of OSI, mean of OSI, slope coefficient of OSI linear regression, coefficient of variation in OSI, interquartile range of OSI, and other 8 parameters were used as model parameters to establish a dacryocystitis screening model with an overall detection accuracy of 85.71%. CONCLUSIONS: This new screening model that is based on ocular surface indicators provides a new option for noninvasive screening of dacryocystitis.

Predictors for Surgeries With the Endoscope-Navigation System for Traumatic Optic Neuropathy and its Clinical Assessment.

BACKGROUND: To assess surgeries with the endoscope-navigation system (ENS) in patients who underwent traumatic optic neuropathy (TON) and find predictors for best corrected visual acuity (BCVA) outcomes. METHODS: The clinical data of 96 consecutive TON patients (96 eyes) who underwent decompression surgery with ENS in the Department of Ophthalmology, Shanghai Ninth People's Hospital, from January 2013 to December 2019 were retrospectively reviewed and analyzed. A binary logistic regression was performed to establish a predictive model for BCVA after treatment as TON outcome. RESULTS: By practicing ENS, 49/96 (51.0%) TON patients got improvement in BCVA, whereas the improvement rate of patients with BCVA of light perception or better was 72.5% (29/40). Hemorrhage within the postethmoid and/or sphenoid sinus, orbital fracture, time interval between trauma and treatment, and BCVA before treatment were predictors for BCVA improvement in TON patients by practicing ENS surgery. The area under raw current curves of the predictive model was 0.826. CONCLUSIONS: Surgeries with the ENS showed positive outcomes for TON patients, especially for those with better BCVA before treatment, shorter time interval between trauma and treatment, without orbital fracture or hemorrhage within the postethmoid and/or sphenoid sinus.

Dacryoendoscopy-assisted incision of Hasner's valve under nasoendoscopy for membranous congenital nasolacrimal duct obstruction after probing failure: a retrospective study.

BACKGROUND: To introduce a treatment option: dacryoendoscopy-assisted incision of Hasner's valve under nasoendoscopy and assess its efficacy in treating membranous congenital nasolacrimal duct obstruction (CNLDO) in children older than 1 year with history of initial probing failure. METHODS: 52 eyes of 43 children with membranous CNLDO who underwent dacryoendoscopy-assisted incision of Hasner's valve under nasoendoscopy between May 2012 and October 2020 were enrolled in this retrospective study. All participants were older than 1 year and all the eyes had gone through probing at least once but still had symptoms of epiphora and discharge. Surgical success was defined as a normal fluorescein dye disappearance test (FDDT) and the absence of pre-operation complaints, including epiphora, muco-purulent discharge, increased tear lake or the symptoms of acute infection such as acute dacryocystitis. Patients' demographics, clinical features and follow-up outcomes were evaluated. RESULTS: Of all these operated cases, surgical success was achieved in 52/52 eyes without any early or late complications. The overall success rate was 100%. CONCLUSIONS: Dacryoendoscopy-assisted incision of Hasner's valve under nasoendoscopy is an effective and minimally invasive surgical treatment in membranous CNLDO patients with initial probing failure.

Eiology and prognosis of canalicular laceration repair using canalicular anastomosis combined with bicanalicular stent intubation.

BACKGROUND: To evaluate the etiology of lacrimal canalicular laceration and explore the possible risk factors influencing prognosis. METHODS: The data of 142 patients (142 eyes) with lacrimal canalicular lacerations who were surgically treated using canalicular anastomosis combined with bicanalicular stent intubation between March 2017 and March 2018 were reviewed. The analyzed data contained demographic information, types of trauma, injury locations, associated additional ocular injuries, and surgical outcomes at follow-up. The main outcome measures were anatomic success rate, functional success rate, and complications of surgery. RESULTS: The mean patient age was 42.07 years (ranging from 1 to 75 years). Among the 142 patients, 112 (78.87%) were males. Upper and lower canalicular lacerations were found in 14 (9.86%) and 112 (78.87%) patients, respectively. Meanwhile, both upper and lower canalicular lacerations were found in 16 (11.27%) patients. Electric bike accidents comprised the leading cause of injury, accounting for 76 (53.52%) cases. There were 100 (70.42%) patients who had lid lacerations without tarsal plate fracture and 42 (29.58%) patients who had lid lacerations with tarsal plate fractures. The anatomic success rate was 98.59% and the functional success rate was 83.8%. The functional reconstruction failure rates were higher in patients with indirect injuries, lid lacerations with tarsal plate fractures, and those with punctum splitting (P < 0.05). Surgical complications were detected in the form of lacrimal punctum ectropion in 3 (2.11%) patients, punctum splitting in 2 (1.41%) patients, and stent extrusion and loss in 2 (1.41%) patients. CONCLUSIONS: Electric bike accidents have become the leading cause of injury instead of motor vehicle accidents because of the changes in the lifestyles of people. Indirect injuries, lid lacerations with tarsal plate fractures, and those with punctum splitting were significantly more likely to lead to poor prognosis, as confirmed by the lower functional success rate of surgery.

Surgical procedure of canaliculoplasty in the treatment of primary canaliculitis associated with canalicular dilatation.

BACKGROUND: Primary canaliculitis is a chronic infection of the proximal lacrimal pathway. We aimed to evaluate surgical outcomes of a canaliculoplasty procedure for primary canaliculitis associated with canalicular dilatation. METHODS: This study enrolled 42 primary canaliculitis patients with canalicular dilatation who underwent canaliculoplasty. All patients were treated with canaliculotomy, curettage of canalicular contents and canaliculoplasty with stent placement. Patients' demographics, clinical features, and follow-up outcomes were evaluated. RESULTS: There were 12 males and 30 females with a mean age of 66.1 ± 13.9 years. The mean duration time from the first onset of signs/symptoms to diagnosis was 30.6 ± 39.5 months. Epiphora (90.5%) and mucopurulent discharge from punctum (85.7%) were the most common signs. Thirty-three out of 42 patients (78.6%) achieved complete remission with a mean follow-up time of 25.3 ± 12.9 months. There were 3 patients found to have canalicular stenosis due to obstruction after surgery. CONCLUSION: Canalicular dilatation is a severe condition of primary canaliculitis, probably due to a combined result of long standing disease and the presence of concretions. The surgical procedure of canaliculoplasty can be a highly effective treatment for primary canaliculitis associated with canalicular dilatation.

The Imbalance of Lymphocyte Subsets and Cytokines: Potential Immunologic Insights Into the Pathogenesis of Chronic Dacryocystitis.

Purpose: To explore the quantitative distributions of different lymphocyte subsets in the lacrimal sac mucosa and identify changes of Th1- and Th2-associated cytokines in the tears of patients with chronic dacryocystitis. Methods: Lacrimal sac mucosal specimens from patients with chronic dacryocystitis were analyzed. Hematoxylin-eosin staining and Masson staining were performed for pathologic analysis, and immunohistochemical staining was performed for the detection of CD3+, CD4+, CD8+, CD20+, Th1, and Th2 lymphocytes. Quantitative real-time PCR was performed to analyze IFN-γ and IL-4 mRNA expression. In addition, tear samples from patients with chronic dacryocystitis and healthy volunteers were collected and analyzed with an antibody array system for Th1- and Th2-associated cytokines and chemokines. Results: Different distribution patterns of lymphocyte subsets were observed in the lacrimal sac walls. Both CD20+ B lymphocytes and CD3+ T lymphocytes accumulated in organized lymphoid follicles, and CD3+ T cells were also distributed in a diffuse manner. Among the two subsets of T cells, CD4+ T cells were more abundant than CD8+ T cells. Both the immunohistochemical staining and real-time PCR results revealed significantly higher expression levels of IFN-γ than those of IL-4. The levels of Th1- and Th2-related cytokines and chemokines measured were significantly higher in the tears of patients than in those of controls. Conclusions: The different distribution patterns of lymphocyte subsets provide insight into a potential immunologic mechanism for dacryocystitis. The cytokines secreted by Th1 or Th2 cells may play a major role in the pathogenesis of dacryocystitis and could be explored as therapeutic targets.

Enhanced bioactivity and osteoinductivity of carboxymethyl chitosan/nanohydroxyapatite/graphene oxide nanocomposites.

Tissue engineering approaches combine a bioscaffold with stem cells to provide biological substitutes that can repair bone defects and eventually improve tissue functions. The prospective bioscaffold should have good osteoinductivity. Surface chemical and roughness modifications are regarded as valuable strategies for developing bioscaffolds because of their positive effects on enhancing osteogenic differentiation. However, the synergistic combination of the two strategies is currently poorly studied. In this work, a nanoengineered scaffold with surface chemistry (oxygen-containing groups) and roughness (R q = 74.1 nm) modifications was fabricated by doping nanohydroxyapatite (nHA), chemically crosslinked graphene oxide (GO) and carboxymethyl chitosan (CMC). The biocompatibility and osteoinductivity of the nanoengineered CMC/nHA/GO scaffold was evaluated in vitro and in vivo, and the osteogenic differentiation mechanism of the nanoengineered scaffold was preliminarily investigated. Our data demonstrated that the enhanced osteoinductivity of CMC/nHA/GO may profit from the surface chemistry and roughness, which benefit the ß1 integrin interactions with the extracellular matrix and activate the FAK-ERK signaling pathway to upregulate the expression of osteogenic special proteins. This study indicates that the nanocomposite scaffold with surface chemistry and roughness modifications could serve as a novel and promising bone substitute for tissue engineering.

CT image-based computer-aided system for orbital prosthesis rehabilitation.

In this paper, a computer-aided system for orbital prosthesis rehabilitation is introduced. With the system, a 3D model of the orbital prosthesis can be easily reconstructed from the CT image of a patient by referring to the normal eye of the patient, and the rehabilitation result by the model can be simulated before the surgery. This facilitates surgeons to design appropriate orbital prosthesis and improve rehabilitation esthetics. Based on the system, the preoperative surgery planning for orbital implant can also be made. This improves the reliability, safety and intuition of the rehabilitation surgery well. The system has been applied to clinical CT images of patients, and the experimental results show effectiveness and acceptability of the system in the clinic.

Repair of orbital bone defects in canines using grafts of enriched autologous bone marrow stromal cells.

BACKGROUND: Bone tissue engineering is a new approach for the repair of orbital defects. The aim of the present study was to explore the feasibility of tissue-engineered bone constructed using bone marrow stromal cells (BMSCs) that were rapidly isolated and concentrated from bone marrow (BM) by the red cell lysis method, then combined with ß-tricalcium phosphate (ß-TCP) to create grafts used to restore orbital bone defects in canines. METHODS: In the experimental group, grafts were constructed using BMSCs obtained by red cell lysis from 20 ml bone marrow, combined with ß-TCP and BM via the custom-made stem cell-scaffold device, then used to repair 10 mm diameter medial orbital wall bony defects in canines. Results were compared with those in groups grafted with BM/ß-TCP or ß-TCP alone, or with defects left untreated as controls. The enrichment of BMSCs and nucleated cells (NCs) in the graft was calculated from the number in untreated bone marrow and in suspensions after red cell lysis. Spiral computed tomography (CT) scans were performed 1, 4, 12 and 24 weeks after implantation in all groups. Gross examination, micro-CT and histological measurements were performed 24 weeks after surgery. The results were analyzed to evaluate the efficacy of bone repair. RESULTS: The number of NCs and of colony-forming units within the scaffolds were increased 54.8 times and 53.4 times, respectively, compared with untreated bone marrow. In the BMSC-BM/ß-TCP group, CT examination revealed that the scaffolds were gradually absorbed and the bony defects were restored. Micro-CT and histological examination confirmed that the implantations led to good repair of the defects, with 6 out 8 orbital defects completely restored in the experimental group, while by contrast, the grafts in the control groups did not fully repair the bony defects, a difference which was statistically significant (p<0.05). CONCLUSIONS: Tissue-engineered bone, constructed using BMSCs isolated by red cell lysis of BM, can restore critical-sized orbital wall defects in canines.

In vitro osteogenic induction of bone marrow stromal cells with encapsulated gene-modified bone marrow stromal cells and in vivo implantation for orbital bone repair.

Osteogenic induction with either growth factors or genetic modification has limitations due to the short half-life and cost of the former, or safety concerns regarding the latter. The objective of this study was to employ a microcapsulation technique to separate genetically modified and nonmodified bone marrow stromal cells (BMSCs) to establish a cost-effective and biosafe osteogenic induction methodology with functional evaluation in vitro and in vivo in a canine model. Autologous BMSCs were isolated and transduced with adenoviral vectors containing either BMP-2 or vascular endothelial growth factor (VEGF) or were dual transduced followed by encapsulation in alginate microcapsules using an electrostatic bead generator. After cocultured with encapsulated cells, normal autologous BMSCs were analyzed for osteogenic differentiation and seeded onto tricalcium phosphate (TCP) scaffolds for in vivo implantation to repair orbital wall bone defects (12 mm in diameter) in a canine model. In vitro assays showed that the expression of the transduced genes was significantly upregulated, with significantly more transduced proteins released from the transduced cells compared with control cells. Importantly, examination of the BMSCs induced by soluble factors released from the encapsulated cells revealed a significant upregulation of expression of osteogenic markers Runx2, BSP, OPN, and OCN in dual-transduction or induction groups. In addition, dual transduction and induction resulted in the highest increase of alkaline phosphatase activity and mineralization compared with other experimental groups. In vivo assays using CT, micro-CT, and histology further supported the qPCR and western blot findings. In conclusion, encapsulation of genetically modified BMSCs was able to release a sufficient amount of BMP-2 and VEGF, which effectively induced osteogenic differentiation of normal-cultured BMSCs and demonstrated bone repair of the orbital wall defect after implantation with ß-TCP in vivo.

Orbital wall repair in canines with beta-tricalcium phosphate and induced bone marrow stromal cells.

Bone tissue engineering is a new approach for the repair of orbital defects. The aim of the present study was to evaluate prefabricated beta-tricalcium phosphate (ß-TCP) combined with autologous bone marrow stromal cells (BMSCs) to repair orbital wall defect in canine models. Defects measuring 10 mm in diameter were created in the orbital medial walls of 12 dogs. The orbits were randomly divided into five groups: group 1, repaired with osteogenesis-induced BMSCs/TCP constructs; group 2, repaired with noninduced BMSCs/TCP constructs; group 3, repaired with ß-TCP scaffolds only; group 4, normal group; group 5, negative control (bone defect without treatment). Computed tomography (CT) scanning, gross observation, bone density measurements, micro-CT, and histological observations were performed. In group 1, new bone was observed with only a small amount of residual material, and bony union was achieved 3 months after surgery. In contrast, the constructs showed slow degradation with minimal bone formation in groups 2 and 3. Furthermore, the appearance and bone density of the constructs in group 1 were similar to that of normal bone: the constructs were covered with complete mucosa, and new alveolate plate grew into the ethmoidal sinuses. A large bone defect remained in group 5. This study demonstrated that biologic scaffolds composed of ß-TCP and osteogenesis-induced BMSCs have been successfully used to restore bone functionality in animal models, which may provide a potential clinical approach for orbital wall repair and bone regeneration in humans.

[A research on three-dimensional reconstruction of nasolacrimal duct in lacrimal surgery].

In this paper, we present a new method which can reconstruct the three-dimensional model of nasolacrimal duct. We firstly resampled the volume data along nasolacrimal duct direction, then segmented the nasolacrimal duct into slices, and finally, completed the 3D reconstruction from the two-D contours. Using this method, we can not only reconstruct normal nasolacrimal duct, but also reconstruct catagmatic nasolacrimal duct. It overcomes the current shortcomings of some traditional methods. Consequently, the technology proposed is of great significance in computer aided technique and surgical planning related to ophthalmonogy.