Enhancing Vault Prediction and ICL Sizing Through Advanced Machine Learning Models.

PURPOSE: To use artificial intelligence (AI) technology to accurately predict vault and Implantable Collamer Lens (ICL) size. METHODS: The methodology focused on enhancing predictive capabilities through the fusion of machine-learning algorithms. Specifically, AdaBoost, Random Forest, Decision Tree, Support Vector Regression, LightGBM, and XGBoost were integrated into a majority-vote model. The performance of each model was evaluated using appropriate metrics such as accuracy, precision, F1-score, and area under the curve (AUC). RESULTS: The majority-vote model exhibited the highest performance among the classification models, with an accuracy of 81.9% area under the curve (AUC) of 0.807. Notably, LightGBM (accuracy = 0.788, AUC = 0.803) and XGBoost (ACC = 0.790, AUC = 0.801) demonstrated competitive results. For the ICL size prediction, the Random Forest model achieved an impressive accuracy of 85.3% (AUC = 0.973), whereas XG-Boost (accuracy = 0.834, AUC = 0.961) and LightGBM (accuracy = 0.816, AUC = 0.961) maintained their compatibility. CONCLUSIONS: This study highlights the potential of diverse machine learning algorithms to enhance postoperative vault and ICL size prediction, ultimately contributing to the safety of ICL implantation procedures. Furthermore, the introduction of the novel majority-vote model demonstrates its capability to combine the advantages of multiple models, yielding superior accuracy. Importantly, this study will empower ophthalmologists to use a precise tool for vault prediction, facilitating informed ICL size selection in clinical practice. [J Refract Surg. 2024;40(3):e126-e132.].

Predictability comparison of sizing parameters for postoperative vault after implantable Collamer lens implantation.

PURPOSE: This study aims to assess the accuracy of three parameters (white-to-white distance [WTW], angle-to-angle [ATA], and sulcus-to-sulcus [STS]) in predicting postoperative vault and to formulate an optimized predictive model. METHODS: In this retrospective study, a cohort of 465 patients (comprising 769 eyes) who underwent the implantation of the V4c implantable Collamer lens with a central port (ICL) for myopia correction was examined. Least absolute shrinkage and selection operator (LASSO) regression and classification models were used to predict postoperative vault. The influences of WTW, ATA, and STS on predicting the postoperative vault and ICL size were analyzed and compared. RESULTS: The dataset was randomly divided into training (80%) and test (20%) sets, with no significant differences observed between them. The screened variables included only seven variables which conferred the largest signal in the model, namely, lens thickness (LT, estimated coefficients for logistic least absolute shrinkage of -0.20), STS (-0.04), size (0.08), flat K (-0.006), anterior chamber depth (0.15), spherical error (-0.006), and cylindrical error (-0.0008). The optimal prediction model depended on STS (R2=0.419, RMSE=0.139), whereas the least effective prediction model relied on WTW (R2=0.395, RMSE=0.142). In the classified prediction models of the vault, classification prediction of the vault based on STS exhibited superior accuracy compared to ATA or WTW. CONCLUSIONS: This study compared the capabilities of WTW, ATA, and STS in predicting postoperative vault, demonstrating that STS exhibits a stronger correlation than the other two parameters.

Comparison of myopic astigmatic correction after cross-assisted SMILE, FS-LASIK, and transPRK.

PURPOSE: To compare astigmatic correction among cross-assisted small-incision lenticule extraction (SMILE), femtosecond laser-assisted in situ keratomileusis (FS-LASIK), and transepithelial photorefractive keratectomy (transPRK). SETTING: The Eye Hospital of Wenzhou Medical University, Zhejiang, China. DESIGN: Prospective comparison study. METHODS: 154 right eyes of 154 patients with astigmatism of -1.00 to -2.75 diopters (D) were included in this study. 64 eyes, 42 eyes, and 48 eyes were receiving SMILE, FS-LASIK, and transPRK, respectively. The SMILE group used cross-axial alignment for head positioning for astigmatism correction. In the FS-LASIK and transPRK groups, static and dynamic cyclotorsion control were used. Changes in ocular parameters and vector analysis were assessed at 6 months postoperatively. RESULTS: The safety and efficacy indices were comparable among the 3 groups at 6 months postoperatively. Residual astigmatism was smallest in the SMILE group (-0.23 ± 0.25 D) compared with that in FS-LASIK (-0.40 ± 0.28 D, P = .009) and transPRK groups (-0.42 ± 0.32 D, P = .001). 53 (82.8%), 36 (85.7%), and 37 (77.1%) eyes achieved an angle of error within ±5 degrees, respectively ( P = .55). Notably, vector analysis showed that the difference vector, the magnitude of the error, and its absolute value were significantly smaller in the SMILE group than those in the other groups ( P < .05). In addition, the higher-order aberrations, especially coma, were significantly induced postoperatively in each group ( P < .001). CONCLUSIONS: Residual astigmatism magnitude was smallest by cross-assisted SMILE, followed by FS-LASIK and transPRK, and the astigmatism axial correction was comparable among groups.

Ethambutol-induced optic neuropathy with rare bilateral asymmetry onset: A case report.

BACKGROUND: Ethambutol-induced optic neuropathy (EON) most commonly manifests as bilateral symmetrical loss of vision and often cause serious and irreversible visual impairment because of the lack of early detection and effective treatment. We followed a case of EON with rare binocular asymmetric clinical manifestations and observed the changes of visual function and retinal structure after drug withdrawal, so as to further understand the clinical characteristics of this disease. CASE SUMMARY: A 54-year-old man complained of gradual visual decline in the left eye. The patient presented with best-corrected visual acuity of 20/20 in the right eye and 20/50 in the left eye. Color vision examination revealed difficulty in reading green color plates in the left eye. The visual field manifested as concentric contraction in the left eye. After nearly a month of drug withdrawal, the right eye had a similar decline in visual function. At the last visit, 19 mo after drug withdrawal, the visual function significantly recovered in both eyes. During follow-up optical coherence tomography (OCT) examination, both eyes manifested the thickness of the retinal nerve fiber layer from mild thickening to thinning and finally temporal atrophy, and the ganglion cell-inner plexiform layer showed significant thinning. The difference was that a reversible structural disorder in the outer retina of the nasal macula was detected in the left eye by macular high-definition OCT. CONCLUSION: Nephropathy and high blood pressure, which damage the retinal microcirculation, may cause damage to the outer layer of the retina. Ethambutol may influence photoreceptor as well as retinal ganglion cells.

Vitrectomy with residual internal limiting membrane covering and autologous blood for a secondary macular hole: A case report.

BACKGROUND: Myopic foveoschisis (MF) is a common complication of pathological myopia. A macular hole (MH) usually results from the natural progression of MF and is a common complication of vitrectomy. Vitrectomy combined with residual internal limiting membrane (ILM) covering and autologous blood was effective for closing a secondary MH. CASE SUMMARY: A 52-year-old woman presented to our clinic with a complaint of blurred vision in the right eye for 7 years. Her best corrected visual acuity (BCVA) was 20/100, axial length was 25.79 mm and standard equivalent refractive error was -10.5 dioptres. Preoperative optical coherence tomography revealed foveoschisis in the right eye. Vitrectomy with fovea-sparing ILM peeling was performed. An MH developed and gradually expanded 5 mo after the initial vitrectomy. Vitrectomy with residual ILM covering and autologous blood was performed. The MH closed 3 wk after the second vitrectomy. CONCLUSION: Fovea-sparing ILM peeling can provide residual ILM for the treatment of MH secondary to vitrectomy for MF. Vitrectomy combined with residual ILM covering and autologous blood is effective for closing secondary MH and improving BCVA.

Bilateral retrocorneal hyaline scrolls secondary to asymptomatic congenital syphilis: A case report.

BACKGROUND: Retrocorneal hyaline scrolls are a rare phenomenon. We report a case of bilateral retrocorneal hyaline scrolls that were likely induced by asymptomatic congenital syphilis. CASE SUMMARY: A 71-year-old woman presented with blurred vision due to cataracts. Slit-lamp microscopy revealed bilateral hyaline scrolls with a dichotomous branching pattern extending to the anterior chamber or rods attaching to the rough posterior surface of the cornea. The patient was positive for syphilis-specific antibodies, with no ocular or systemic evidence of congenital or acquired syphilis. Binocular cataract, retrocorneal scroll, and corneal endothelial gutta were considered. The scroll of the right eye was removed during cataract surgery and further observed using hematoxylin-eosin staining and scanning electron microscopy. The cornea of the right eye remained transparent, and the residual scroll seemed stable, however, the corneal endothelial density declined at 13 mo after surgery. In vivo confocal microscopy revealed coalescence of corneal guttae at the level of the corneal endothelium or adhesion to the posterior surface of the endothelium, with enlarged endothelial cells in both eyes. Activated keratocytes in the stroma and a highly reflective acellular structure at the level of the Descemet's membrane were observed. The removed scroll had a cartilage-like hardness and a circularly arranged fiber-like acellular structure. CONCLUSION: Occult congenital syphilis could induce corneal endothelial gutta and the formation of retrocorneal scrolls without other signs of ocular syphilis.

Analysis of structural injury patterns in peripapillary retinal nerve fibre layer and retinal ganglion cell layer in ethambutol-induced optic neuropathy.

BACKGROUND: We investigated structural injury patterns in the peripapillary retinal nerve fibre layer (p-RNFL) and ganglion cell inner plexiform layer (GCIPL) caused by ethambutol treatment. METHODS: Sixty-four patients undergoing ethambutol treatment at Zhejiang Chinese Medicine and Western Medicine Integrated Hospital were recruited. Fourteen (14) exhibited visual dysfunction (abnormal group), and the remaining 50 had no visual dysfunction (subclinical group). The thickness of the p-RNFL, total macular retina layer and GCIPL were measured using Cirrus-HD Optical coherence tomography (Cirrus-HD OCT, Cirrus high-definition optical coherence tomography), and compared with 60 healthy, age-matched controls. RESULTS: The p-RNFL thickness was similar in both subclinical and control groups. When compared with the control group, p-RNFL thickness in the abnormal group was significantly increased in the inferior and superior quadrants (GEE, P = 0.040, P = 0.010 respectively). In contrast with the subclinical group, p-RNFL thickness in the inferior quadrant was increased in the abnormal group (GEE, P = 0.047). The GCIPL thickness in the inferonasal and inferior sectors was significantly deceased in the subclinical group when compared with controls (GEE, P = 0.028, P = 0.047, respectively). The average and minimum value of GCIPL thickness, and thickness in the superonasal, inferior, inferotemporal, superotemporal and superior sectors were significantly decreased in the abnormal group when compared with controls (GEE, P = 0.016, P = 0.001, P = 0.028, P = 0.010, P = 0.012, P = 0.015, P = 0.010, respectively). The cube average macular thickness (CAMT) in the abnormal group was significantly thinner than controls (GEE, P = 0.027). CONCLUSIONS: GCIPL measurements using Cirrus-HD OCT detected retinal ganglion cell layer loss following ethambutol treatment, before visual dysfunction occurred.

Renal cell carcinoma choroidal metastasis recorded by smartphone with interface eyepiece adapter mounted on slit lamp: A case report.

RATIONALE: Ocular metastasis of renal cell carcinoma (RCC) is rare, and mainly located on the choroid. We report a choroidal metastasis from RCC, which was recorded by a smartphone with an interface eyepiece adapter mounted on a slit lamp. PATIENT CONCERNS: A 45-year-old female presented with 1-month history of painless occlusion of the vision field on the left eye, who had undergone right nephrectomy for RCC 19 months ago. DIAGNOSES: A smooth, hemispherical and brown protrusion was found behind the pupil nasally. An enhanced computed tomography scan of the orbit showed a slightly high-density hemispherical nodule involving the nasal portions of the left eyeball, the enhancement of the lesion was obvious and homogeneous. A metastatic choroidal space-occupying lesion from RCC was highly suspected according to the clinical and radiological findings. INTERVENTIONS: The patient was advised to undergo further treatment, such as radiotherapy. OUTCOMES: The images of choroid metastasis were recorded by the smartphone with the interface eyepiece adapter mounted on the slit lamp handily. CONCLUSIONS: The smartphone with an interface eyepiece adapter mounted on the slit lamp can be widely used to record the precious images in the clinic in a timely manner.

A novel role for Livin in the response to ultraviolet B radiation and pterygium development.

A pterygium is an inflammatory, invasive and proliferative lesion on the ocular surface, which can decrease visual acuity, damage the ocular surface and affect the appearance of the eye. However, the underlying molecular mechanisms of the pathogenesis remain unclear. In the present study, the role of apoptosis­associated protein Livin in the occurrence and development of pterygium was investigated. Primary samples from quiescent or advanced clinical stages of pterygium and normal human conjunctival tissues were used to assess mRNA and protein expression levels of Livin using reverse transcription­quantitative PCR and immunohistochemistry, respectively. Livin was knocked down in pterygium epithelial cells (PECs) using small interfering RNA (siRNA), to investigate the role of Livin in PEC viability, migration, invasion ability and apoptosis. The cell viability, invasion ability and apoptosis of PECs following ultraviolet B (UVB) radiation alone or in combination with Livin silencing were also analyzed. Expression levels of Livin increased in the pterygium tissues compared with those in the normal conjunctiva at both the mRNA and protein levels. Livin expression levels in advanced pterygium were significantly higher compared with those in quiescent pterygium samples. Knockdown of Livin expression levels significantly reduced cell migration, invasion ability and cell viability, and induced apoptosis of PECs. Inhibition of Livin expression in PECs increased the expression levels of caspase­7, caspase­3 and E­cadherin, whereas expression levels of Snail were downregulated. Cell viability and invasion ability in PECs was enhanced following UVB radiation and Livin expression upregulated. UVB irradiation induced cell invasion ability of PECs and this was attenuated by Livin­silencing. Transfection with Livin siRNA also partially recovered the apoptosis rate of PECs, which was reduced by UVB irradiation. In conclusion, Livin was upregulated in pterygium, and UVB radiation functions in the development of pterygium by inducing Livin expression.

Spontaneous rupture of the eyeball due to choroidal metastasis of gastric carcinoma: A case report.

RATIONALE: Although metastatic tumor is the most common intraocular malignancy, choroidal metastasis from gastric cancer is relatively rare. We present the exact course of a spontaneous rupture of the eyeball with choroidal metastasis from gastric carcinoma (GC) and the applied surgical interventions. PATIENT CONCERNS: A 59-year-old male presented with pain and vision loss on his left eye for 6 months. He was diagnosed with GC, for which he received systemic chemotherapy for a year. DIAGNOSES: Preoperative B-scan, color fundus photography, computed tomography, and magnetic resonance imaging showed a choroidal tumor in his left eye. The spontaneous rupture of the corneoscleral limbus from 2 to 5 o'clock, combined secondary glaucoma, exudative retinal detachment and choroidal detachment were found. INTERVENTIONS: The ruptured corneoscleral limbus was sutured and the orbit was lavaged with 0.4% cisplatin during the enucleation. OUTCOMES: Histopathology confirmed high homology of the choroidal metastasis and GC. He survived for 2 months after surgery, without pain or orbital neoplasms. LESSONS: Choroidal metastasis from GC rapidly progressed to spontaneous rupture of the eyeball. Careful eyeball enucleation followed by orbital lavage with chemotherapeutics may reduce metastasis risk beyond the eyeball. Additional therapeutic interventions should be considered in patients resistant to single systemic chemotherapy.

Unilateral inferior oblique anterior transposition for markedly asymmetric dissociated vertical deviation with unilateral inferior oblique over-action.

BACKGROUND: To evaluate the results of unilateral inferior oblique anterior transposition (IOAT) for markedly asymmetric dissociated vertical deviation (DVD) combined with inferior oblique over-action (IOOA). METHODS: Retrospective chart review of the records of all patients with asymmetric DVD combined with unilateral IOOA in the non-dominant eye who received unilateral IOAT on the non-dominant eye. No other muscles were operated on simultaneously. The amount of DVD and IOOA were measured before and after the operation and statistically analysed. RESULTS: Seventeen patients were included. The mean age at surgery was 23.5 ± 8.4 (range 12-38) years old. The mean postoperative follow-up period was 15.7 ± 7.2 (range 6-32) months. The primary position DVD was 19.6 ± 5.4 (range 14-36) PD preoperatively and decreased significantly to 2.9 ± 2.0 (range 0-8) PD postoperatively (P < 0.01). Preoperatively, there were 2, 7, and 8 patients with + 1, + 2, and + 3 IOOA, respectively, and these were reduced from 2.4 ± 0.7 to 0.3 ± 0.4 postoperatively (P < 0.01). None of the patients were complicated obvious hypotropia, anti-elevation syndrome or IOOA in the contralateral eye. CONCLUSIONS: Unilateral IOAT was recommended in patients with asymmetric DVD coexists with unilateral IOOA.

Long-term comparison of full-bed deep anterior lamellar keratoplasty and penetrating keratoplasty in treating keratoconus.

OBJECTIVE: To compare postoperative outcomes of full-bed deep anterior lamellar keratoplasty (DALK) with penetrating keratoplasty (PK) in treating keratoconus. METHODS: Seventy-five eyes of 64 patients who received full-bed DALK and 52 eyes of 51 patients who received PK between June 2000 and August 2010 were included in this retrospective study. Full-bed DALK was performed using Yao's hooking-detaching technique. PK was performed using a standard technique. Intraoperative and postoperative complications, visual acuity, rejection, graft survival, endothelial cell density, corneal sensation recovery, and re-innervation were compared between the two groups. RESULTS: A best correct visual acuity of 0.5 or better was achieved in 90.7% of eyes after full-bed DALK and in 92.3% of eyes after PK (P=0.75). By the fifth postoperative year, graft endothelial cell loss reached 34.6% in the PK group vs. 13.9% in the full-bed DALK group (P<0.001). There were no statistical differences in corneal sensitivity recovery or corneal re-innervation between the groups (P>0.05). Intraoperative microperforation occurred in seven out of 75 (9.3%) eyes with a temporally postoperative double anterior chamber in two eyes in the full-bed DALK group. Postoperative complications in the PK vs. the full-bed DALK groups respectively were: rejection (7.7% vs. 0%, P=0.015), high intraocular pressure (IOP) (46.2% vs. 1.3%, P<0.001), secondary glaucoma (9.6% vs. 0%, P=0.006), complicated cataract (19.2% vs. 0%, P<0.001), and wound dehiscence (9.6% vs. 0%, P=0.006). CONCLUSIONS: Both full-bed DALK and PK can offer long-term satisfactory visual outcomes for keratoconus. Graft rejection, secondary glaucoma, complicated cataracts, and constant endothelial cell loss were observed in eyes only after PK.

Macular and peripapillary retinal nerve fiber layer thickness in children with hyperopic anisometropic amblyopia.

AIM: To compare the retinal nerve fiber layer (RNFL) thickness and macular thickness in the amblyopic eye with that in the sound eye of children with hyperopic anisometropic amblyopia using optical coherence tomography (OCT). METHODS: A prospective, nonrandom, intraindividual comparative cohort study includes 72 children with hyperopic anisometropic amblyopia in a single center. Macular thickness, macular foveola thickness, and peripapillary RNFL thickness were compared between the amblyopia eyes and the contralateral sound eyes. RESULTS: There were 38 male and 34 female patients, with a mean age as 9.7±1.9 years (range, 5-16 years). Hyperopic was +3.62±1.16D (range +2.00D to +6.50D) in the amblyopic eyes, which was significantly higher in the control eyes with +0.76±0.90D (range 0D to +2.00D) (P < 0.01). The mean peripapillary RNFL thickness was 113.9±7.2µm and 109.2±6.9µm in the amblyopic eye and the normal eye, respectively, reaching statistical significance (P = 0.02). The mean macular foveola thickness was significantly thicker in the amblyopic eyes than the contralateral sound eyes (181.4±14.2µm vs 175.2±13.3µm, P < 0.01), but the 1mm, 3mm or 6mm macular thickness central macular thickness was not significantly different. Degree of anisometropia in the contralateral eyes was not significantly correlated with differences of peripapillary RNFL, macular foveola thickness or central macular thickness. CONCLUSION: Eyes with hyperopic anisometropic amblyopia are found thicker macular foveola and peripapillary RNFL than the contralateral eyes in children.

[Study on the chemical constituents from Gnaphalium hypoleucum].

OBJECTIVE: To research the chemical constituents from Gnaphalium hypoleucum. METHODS: Compounds were separated by using silica gel, ODS, Sephadex LH-20 chromatography and their structures were elucidated by means of spectral data analysis. RESULTS: Ten compounds were isolated and identified as tetracosanoic acid (1), 5-hydroxy-3,6,7,8-tetramethoxyflavone (2), beta-sitosterol (3), 5-hydroxy-3,6,7,8,4'-pentamethoxyflavone (4), 5-hydroxy-3,6, 7,8,3',4'-hexamethoxyflavone (5), apigenin (6), luteolin (7), quercetin (8), luteolin4'-O-beta-D-glucoside (9), quercetin-4'-O-beta-D-glucoside (10). CONCLUSION: All of these compounds are isolated from this plant for the first time. Compound 5 is isolated from the genus for the first time.

Three new acyclic diterpenoids from Eupatorium lindleyanum DC.

Three new acyclic diterpenoids were isolated from the whole plant of Eupatorium lindleyanum DC. The structures of the compounds were elucidated by means of (1)H and (13)C NMR spectroscopic analyses, including 2D NMR experiments.

Long-term comparison of full-bed deep lamellar keratoplasty with penetrating keratoplasty in treating corneal leucoma caused by herpes simplex keratitis.

PURPOSE: To compare long-term outcomes of full-bed deep lamellar keratoplasty (DLK) with penetrating keratoplasty (PK) for treating corneal leucoma caused by herpes simplex keratitis (HSK). DESIGN: Retrospective, comparative, interventional case series. METHODS: setting: Institutional. patients: Inclusion criteria were patients with corneal scarring induced exclusively by HSK who underwent primary graft of full-bed DLK or PK and completed a minimum of 12 months postoperative follow-up. There was no significant difference of corneal scarring and vascularization between the 2 groups before surgery. Choosing PK or full-bed DLK depended on the patient's own willingness, except those patients with a preoperative endothelial cell count of less than 700 cells/mm(2) or whose endothelial cell count was undetectable were encouraged to undergo only PK. Exclusion criteria were patients with a past history of corneal perforation, nonprimary graft, non-HSK-related corneal scars, and failure to complete a minimum of 12 months of postoperative follow-up. Fifty-eight eyes of 58 patients in the full-bed DLK group and 63 eyes of 63 patients in the PK group met the inclusion criteria. main outcome measures: Postoperative managements, recurrence of HSK, graft rejection, graft survival rate, visual acuity, and corneal endothelial density. RESULTS: The mean postoperative follow-up duration was 45.8 ± 30.9 months in the full-bed DLK group and 47.9 ± 27.2 months in the PK group (P = .70). As compared with the PK group, the full-bed DLK group experienced earlier suture removal (P = .01), needed fewer postoperative visits (P < .001), and had a higher proportion of eyes with full withdrawal of oral acyclovir (P < .001) and topical corticosteroid (P < .001). There were a total of 21 episodes of recurrent HSK in the PK group, more frequent than the 7 episodes in the full-bed DLK group, among which recurrent epithelial keratitis amounted to 13 episodes in the PK group, remarkably more frequent than the 1 episode in the full-bed DLK group. Twenty-six eyes (41.3%) encountered rejection episodes in the PK group, but no rejection episode was found in the full-bed DLK group (P < .001). In 14 eyes in the PK group, graft failure developed because of graft rejection, recurrence of HSK, or both, whereas only in 1 eye in the full-bed DLK group did graft failure develop because of recurrence of HSK (P = .001). The clear graft survival rate in the full-bed DLK group was significantly higher than that in the PK group (P = .01). Corneal endothelial cell density was stable from 1 month through 5 years in the full-bed DLK group, but 51.3% cell loss was found in the PK group at 5 years after surgery. At the last visit, 66.1% of eyes with full-bed DLK grafts and 50.9% of eyes with PK grafts achieved a best-correct visual acuity of 0.5 or better (P = .10). CONCLUSIONS: Advantages of full-bed DLK over PK are no allograft rejection, longer graft survival, earlier drug withdrawal of topical steroid and oral acyclovir, less recurrence of HSK, and fewer follow-up visits. Full-bed DLK is preferable for treating HSK-induced corneal scarring with relatively healthy endothelium and with no history of perforation.

General nonextremal rotating charged Gödel black holes in minimal five-dimensional gauged supergravity.

I present the general exact solutions for nonextremal rotating charged black holes in the Gödel universe of five-dimensional minimal supergravity theory. They are uniquely characterized by four nontrivial parameters: namely, the mass m, the charge q, the Kerr equal rotation parameter a, and the Gödel parameter j. I calculate the conserved energy, angular momenta, and charge for the solutions and show that they completely satisfy the first law of black hole thermodynamics. I also study the symmetry and separability of the Hamilton-Jacobi and the massive Klein-Gordon equations in these Einstein-Maxwell-Chern-Simons-Gödel black hole backgrounds.

[Expression of recombinant human BMP-6 in Escherichia coli and its purification and bioassay in vitro].

To purify the recombinant human BMP-6 protein and to establish its in vitro bioassay method. The cDNA encoding the mature peptide of hBMP-6 protein was amplified by reverse transcription-polymerase chain reaction (RT-PCR), using human placental mRNA as template, and subcloned into the high-expression vector pET-15b under the control of T7 lac promoter. The resulting construct, pET-BMP6, was then transformed into an Escherichia coli strain BL21 (DE3) for the production of recombinant hBMP-6 protein (rhBMP-6). After 4 hours of induction by isopropyl-beta-D-thiogalactoside (IPTG), rhBMP-6 (approximately 15kD) was expressed and formed inclusion bodies, contributing up to 10% of the total bacterial protein. The inclusion bodies were isolated and redissolved in 8mol/L urea, and the denatured rhBMP-6 was purified to 95% purity by CM-Cellulose 32 ion exchange chromatography (IEC). The osteoinductivity of rhBMP-6 was measured by the expression of some of the osteoblast differentiation marker genes in rhBMP-6-treated C3H10T1/2 cells as reflected by determinations of alkaline phosphatase (ALPase) activity and semi-quantitative RT-PCR. At the end of the purification process, about 80% of rhBMP-6 formed disulphide-linked homodimers after refolding during renaturation. The apparent size of the protein was 30kD on non-reducing SDS-PAGE, similar to that of the native form of hBMP-6. The enzyme assays showed that the ALPase activity was increased in a dose-dependent manner with the treatment of rhBMP-6. After the addition of 300ng/mL of rhBMP-6, the ALPase activity of C3H10T1/2 cells increased significantly. The activity of rhBMP-6 used was comparable to about 70% of that of the standard hBMP-6 derived from eukaryotic cells. RNA extraction data also showed rhBMP-6 stimulated expression of osteoblast marker genes, including type I collagen, osterix, and osteocalcin in a time-dependent manner. After 5 days of treatment, their level of expression was increased to 3 times that of controls. Bone morphogenetic protein (BMP)-6, a member of the 60A subgroup of the bone morphogenetic protein (BMPs) family, plays a pivotal role in bone formation. Previous evidence showed that BMP-6 is selectively up-regulated by estrogen, suggesting its potential role in the treatment of osteoporotic fractures, especially for menopausal osteoporosis. Our present study demonstrates that the recombinant hBMP-6 produced in Escherichia coli is able to induce pre-osteoblastic cells to differentiate into osteoblasts in vitro, and analysis of mRNA expression of type I collagen, osterix, and osteocalcin can be also a method for measuring the osteoinductivity of BMP. This provides the basis for further studies on ectopic bone formation in the body and for the development of auxiliary drugs for the treatment of osteoporotic fractures.