Androgen synthesis cell-specific CREBZF deficiency alters adrenal cortex steroid secretion and develops behavioral abnormalities in adult male mice.

The global challenge of male infertility is escalating, notably due to the decreased testosterone (T) synthesis in testicular Leydig cells under stress, underscoring the critical need for a more profound understanding of its regulatory mechanisms. CREBZF, a novel basic region-leucine zipper transcription factor, regulates testosterone synthesis in mouse Leydig cells in vitro; however, further validation through in vivo experiments is essential. Our study utilized Cyp17a1-Cre to knock out CREBZF in androgen-synthesis cells and explored the physiological roles of CREBZF in fertility, steroid hormone synthesis, and behaviors in adult male mice. Conditional knockout (cKO) CREBZF did not affect fertility and serum testosterone level in male mice. Primary Leydig cells isolated from CREBZF-cKO mice showed impaired testosterone secretion and decreased mRNA levels of Star, Cyp17a1, and Hsd3b1. Loss of CREBZF resulted in thickening of the adrenal cortex, especially X-zone, with elevated serum corticosterone and dehydroepiandrosterone levels and decreased serum dehydroepiandrosterone sulfate levels. Immunohistochemical staining revealed increased expression of StAR, Cyp11a1, and 17ß-Hsd3 in the adrenal cortex of CREBZF-cKO mice, while the expression of AR was significantly reduced. Along with the histological changes and abnormal steroid levels in the adrenal gland, CREBZF-cKO mice showed higher anxiety-like behavior and impaired memory in the elevated plus maze and Barnes maze, respectively. In summary, CREBZF is dispensable for fertility, and CREBZF deficiency in Leydig cells promotes adrenal function in adult male mice. These results shed light on the requirement of CREBZF for fertility, adrenal steroid synthesis, and stress response in adult male mice, and contribute to understanding the crosstalk between testes and adrenal glands.

Economic Evaluation of COVID-19 Immunization Strategies: A Systematic Review and Narrative Synthesis.

OBJECTIVES: This study aimed to systematically assess global economic evaluation studies on COVID-19 vaccination, offer valuable insights for future economic evaluations, and assist policymakers in making evidence-based decisions regarding the implementation of COVID-19 vaccination. METHODS: Searches were performed from January 2020 to September 2023 across seven English databases (PubMed, Web of Science, MEDLINE, EBSCO, KCL-Korean Journal Dataset, SciELO Citation Index, and Derwent Innovations Index) and three Chinese databases (Wanfang Data, China Science and Technology Journal, and CNKI). Rigorous inclusion and exclusion criteria were applied. Data were extracted from eligible studies using a standardized data collection form, with the reporting quality of these studies assessed using the Consolidated Health Economic Evaluation Reporting Standards 2022 (CHEERS 2022). RESULTS: Of the 40 studies included in the final review, the overall reporting quality was good, evidenced by a mean score of 22.6 (ranging from 10.5 to 28). Given the significant heterogeneity in fundamental aspects among the studies reviewed, a narrative synthesis was conducted. Most of these studies adopted a health system or societal perspective. They predominantly utilized a composite model, merging dynamic and static methods, within short to medium-term time horizons to simulate various vaccination strategies. The research strategies varied among studies, investigating different doses, dosages, brands, mechanisms, efficacies, vaccination coverage rates, deployment speeds, and priority target groups. Three pivotal parameters notably influenced the evaluation results: the vaccine's effectiveness, its cost, and the basic reproductive number (R0). Despite variations in model structures, baseline parameters, and assumptions utilized, all studies identified a general trend that COVID-19 vaccination is cost-effective compared to no vaccination or intervention. CONCLUSIONS: The current review confirmed that COVID-19 vaccination is a cost-effective alternative in preventing and controlling COVID-19. In addition, it highlights the profound impact of variables such as dose size, target population, vaccine efficacy, speed of vaccination, and diversity of vaccine brands and mechanisms on cost effectiveness, and also proposes practical and effective strategies for improving COVID-19 vaccination campaigns from the perspective of economic evaluation.

LncRNA STAT3-AS regulates endometrial receptivity via the STAT3 signaling pathway.

Endometrial receptivity is critical for the successful establishment of pregnancy in ruminants. Interferon tau (IFNT) plays a key role in promoting embryo attachment by activating the Janus kinase/signal transducer and activator of transcription pathway, which induces the expression of a series of interferon-stimulated genes (ISGs). In our previous study, sequencing analysis of goat endometrial epithelial cells (gEECs) treated with 20 ng/mL IFNT revealed a differentially expressed long non-coding RNA located on the STAT3 antisense chain, which we designated STAT3-AS. The aim of this study was to investigate the role and mechanism of STAT3-AS in establishing endometrial receptivity in goats. The results showed that STAT3-AS was expressed in both the nucleus and cytoplasm of gEECs, and its expression increased significantly in the uterus on day 15 of pregnancy. STAT3-AS expression was upregulated in gEECs treated with IFNT alone or in combination with progesterone and estradiol. Knockdown of STAT3-AS using specific short interfering RNA significantly inhibited the expression of classical ISGs such as interferon-stimulated gene 15 and 2',5'-oligodenylate synthetase 2, as well as uterine endometrial receptivity-related genes including homeobox gene A11, integrin beta 3, and vascular endothelial growth factor. Moreover, gEEC proliferation and the STAT3 pathway were suppressed in the absence of STAT3-AS. However, pretreatment with the STAT3 activator RO8191 restored the effect of silencing STAT3-AS on endometrial receptivity. Overall, these results suggest that STAT3-AS is an important regulator of endometrial receptivity in goats and that it regulates endometrial receptivity through the STAT3 pathway.

A head-to-head comparison of the measurement properties of EQ-5D-3L and EQ-5D-5L in Chinese family caregivers of cancer patients.

BACKGROUND: Although both EQ-5D-3L(3L) and EQ-5D-5L(5L) have demonstrated good measurement properties in several patient populations, there is currently limited evidence comparing the measurement properties of 3L and 5L in family caregivers (FCs) of cancer patients. PURPOSE: This study aimed to compare the measurement properties of 3L and 5L in a sample of family caregivers of cancer patients. METHODS: A consecutive sample of FCs of cancer patients recruited from three tertiary hospitals were invited to complete the two versions of the EQ-5D in two rounds of interviews. We compared i) the ceiling effect using the McNemar's test, ii) test-retest reliability using intraclass correlation coefficient (ICC) and Cohen's Kappa, iii) convergent validity using Spearman's rank correlation coefficient, iv) known-group validity using F-statistic, v) and discriminant capacity using ordinal logistic regression. RESULTS: A total of 416 FCs completed the baseline questionnaire and 120 caregivers completed the follow-up questionnaire. Ceiling effects were smaller in 5L (12.5%) than in 3L (20.7%). The convergent validity (r = 0.344-0.771), known-groups validity (Fratio5L/3L = 2.06-4.09), discriminant capacity (ES = 0.341-0.396), and test-retest reliability (ICC = 0.725) of the 5L were slightly better than those of the 3L in China. CONCLUSION: The current study found both 3L and 5L to be suitable for use by FCs of cancer patients. However, 5L showed superior measurement properties compared to 3L and therefore could be the preferred instrument when EQ-5D data of cancer patients FCs is required.

USP18 promotes endometrial receptivity via the JAK/STAT1 and the ISGylation pathway.

Interferon-tau (IFNT), a pregnancy recognition signal in ruminants, promotes the establishment of endometrial receptivity by inducing the expression of interferon-stimulated genes (ISGs) via the Janus kinase/signal transducer and activator of transcription (JAK/STATs) signaling pathway. However, the precise mechanisms remain largely unknown. Ubiquitin-specific protease 18 (USP18) acts specifically on the ISGylation modification system to exert deubiquitination and participates in the regulation of the type I IFN signaling pathway. The purpose of this study was to determine the role and mechanism of USP18 on endometrial receptivity in goat. USP18 was mainly localized in the uterine luminal and glandular epithelium, and its expression levels were significantly increased from days 5-18 of early pregnancy. Progesterone (P4), estradiol (E2), and IFNT significantly stimulated USP18 expression in goat endometrial epithelial cells (gEECs) cultured in vitro. Meanwhile, the markers of endometrial receptivity HOXA11, ITGB1, ITGB3, and ITGB5 were significantly upregulated after USP18 overexpression in gEECs. However, USP18 interference significantly inhibited the expression of HOXA10, ITGB1, ITGB3, and ITGB5 in gEECs. In addition, both the phosphorylation levels of STAT1 and the expression of ISGylation-modified proteins were significantly increased after USP18 silencing in gEECs. Furthermore, pretreatment with the STAT1 inhibitor Fludara markedly restored the effect of USP18 interference in gEECs. In summary, USP18 may play an important role in promoting goat endometrial receptivity by regulating the JAK/STAT1 pathway and ISGylation.

Hand2os1 Regulates the Secretion of Progesterone in Mice Corpus Luteum.

The corpus luteum plays a key role in pregnancy maintenance and estrous cycle regulation by secreting progesterone. Hand2os1 is an lncRNA located upstream of Hand2, with which a bidirectional promoter is shared and is involved in the regulation of cardiac development and embryo implantation in mice. The aim of this study was to investigate the expression and regulation of Hand2os1 in the ovaries. Here, we used RNAscope to detect differential expression of Hand2os1 in the ovaries of cycling and pregnant mice. Hand2os1 was specifically detected in luteal cells during the proestrus and estrus phases, showing its highest expression in the corpus luteum at estrus. Additionally, Hand2os1 was strongly expressed in the corpus luteum on day 4 of pregnancy, but the positive signal progressively disappeared after day 8, was detected again on day 18, and gradually decreased after delivery. Hand2os1 significantly promoted the synthesis of progesterone and the expression of StAR and Cyp11a1. The decreased progesterone levels caused by Hand2os1 interference were rescued by the overexpression of StAR. Our findings suggest that Hand2os1 may regulate the secretion of progesterone in the mouse corpus luteum by affecting the key rate-limiting enzyme StAR, which may have an impact on the maintenance of pregnancy.

Long-term corneal recovery by simultaneous delivery of hPSC-derived corneal endothelial precursors and nicotinamide.

Human pluripotent stem cells (hPSCs) hold great promise for the treatment of various human diseases. However, their therapeutic benefits and mechanisms for treating corneal endothelial dysfunction remain undefined. Here, we developed a therapeutic regimen consisting of the combination of hPSC-derived corneal endothelial precursors (CEPs) with nicotinamide (NAM) for effective treatment of corneal endothelial dysfunction. In rabbit and nonhuman primate models, intracameral injection of CEPs and NAM achieved long-term recovery of corneal clarity and thickness, similar with the therapeutic outcome of cultured human corneal endothelial cells (CECs). The transplanted human CEPs exhibited structural and functional integration with host resident CECs. However, the long-term recovery relied on the stimulation of endogenous endothelial regeneration in rabbits, but predominantly on the replacing function of transplanted cells during the 3-year follow-up in nonhuman primates, which resemble human corneal endothelium with limited regenerative capacity. Mechanistically, NAM ensured in vivo proper maturation of transplanted CEPs into functional CECs by preventing premature senescence and endothelial-mesenchymal transition within the TGF-ß-enriched aqueous humor. Together, we provide compelling experimental evidence and mechanistic insights of simultaneous delivery of CEPs and NAM as a potential approach for treating corneal endothelial dysfunction.

Ufmylation regulates granulosa cell apoptosis via ER stress but not oxidative stress during goat follicular atresia.

Follicular atresia is primarily caused by granulosa cell (GC) apoptosis, although the mechanisms are largely unknown. Ufmylation is a recently identified ubiquitin-like post-translational modifier that plays an important role in cell proliferation and apoptosis. The purpose of this study was to investigate the effects of Ufmylation on GC apoptosis during goat follicular atresia. Ubiquitin-fold modifier 1 (UFM1) and its target DDRGK domain containing 1 (DDRGK1) proteins were identified in granulosa cells (GCs) isolated from all stages of preantral follicles and from healthy (HF), early atretic (EF) and progressed atretic (PF) antral follicles. The expression levels were higher in GCs derived from antral atretic follicles than healthy follicles. Although the viability of GCs was not affected after overexpression of UFM1, siRNA-mediated UFM1 silencing significantly inhibited GC proliferation and induced apoptosis. Notably, components of the ufmylation pathway were significantly upregulated in GCs induced by the ER stress agent tunicamycin (Tm) and thapsigargin (Tg), but not affected by oxidative stress inducer H2O2. Furthermore, UFM1 silencing markedly increased the apoptosis of GCs upon Tg treatment by stimulating the ER stress-related gene expression. Our results provide evidence that UFM1 and its target DDRGK1 are expressed in the goat GCs during follicular development and atresia, and ufmylation may play an important role in the prevention of ER stress but not oxidative stress-induced GCs apoptosis.

Clinical outcomes of double continuous suture in femtosecond laser-assisted lamellar keratoplasty for keratoconus.

To assess the surgical outcomes in patients who underwent femtosecond laser-assisted lamellar keratoplasty with double continuous suture for keratoconus, 100 patients (102 eyes) with keratoconus in advanced stages undergoing femtosecond laser-assisted lamellar keratoplasty in Shandong Eye Hospital were studied. In the management of keratoconus, 50 patients (52 eyes) received double continuous suture, and 50 patients (50 eyes) underwent interrupted suture. The follow-up duration was 1 year. Best-corrected visual acuity (BCVA), corneal astigmatism, cosmetic outcomes, and surgical complications were measured as outcome indicators. The epithelium healed at 3 ± 2 days and 4 ± 2 days in the double continuous suture group and the interrupted suture groups, respectively (P > 0.05). At 6 months after surgery, the average visual acuity was 20/125 and 20/100 (P > 0.05), and the average BCVA was 20/32 and 20/40 (P > 0.05), respectively. At 1 year after surgery, the average visual acuity was 20/63 and 20/80 (P > 0.05), and the average BCVA was 20/32 and 20/25 (P > 0.05), respectively; the mean curvature was 43.24 ± 5.15 D and 43.31 ± 5.58 D (P > 0.05), the mean astigmatism was 3.21 ± 1.74 D and 5.35 ± 1.37 D (P < 0.05).The looseness of sutures were found in 2 patients and 15 patients in both groups, respectively (P < 0.05). No postoperative infection or immune rejection occurred in either group during the follow-up. Comparing with the interrupted suture, using the continuous suture in femtosecond laser-assisted lamellar keratoplasty for keratoconus markedly limited the looseness of sutures with lesser corneal astigmatism and better visual quality postoperative.

Individualized Corneal Patching for Treatment of Corneal Trauma Combined with Tissue Defects.

AIM: To evaluate the efficacy of individualized corneal patching using a minimal graft for corneal trauma combined with tissue defects. METHODS: Fifteen eyes (15 patients) were enrolled in this study, including 8 eyes with corneal perforation induced by removal of metal foreign bodies, 5 eyes with corneal laceration resulting from metal trauma, and 2 eyes with pencil injuries to the cornea. The size, shape, and depth of the tissue defects were assessed. For corneal perforation or irregular tissue defects, if the diameter or length was ≥3.0 mm, traditional penetrating keratoplasty (PK) or lamellar keratoplasty (LK) was adopted; if the diameter or length was ＜3.0 mm, a conical or irregular patch consistent with the defects was used. The visual acuity, corneal status, and postoperative complications were observed during the follow-up. RESULTS: The diameter of corneal perforations was 1.0 mm in 2 eyes, 1.5 mm in 1 eye, 2.0 mm in 4 eyes, and 3.5 mm in 1 eye. During their PK procedures, a conical corneal graft was used in 7 eyes, while a traditional cylindrical graft was used in 1 eye. The other 7 eyes had corneal trauma combined with irregular tissue defects, which were full-thickness corneal defects in 5 eyes and lamellar defects in 2 eyes, all less than 3.0 mm in length. Thus, five eyes received PK, and 2 eyes received LK using an irregular wedge-shaped patch. The visual acuity increased greatly postoperatively, with mild corneal astigmatism. None of the patients developed immune rejection. CONCLUSION: Individualized corneal patching with a minimal graft can save corneal materials, relieve corneal scars, gain a good visual prognosis, and avoid immune rejection in the treatment of corneal trauma combined with tissue defects.

UFMylation is associated with LPS-induced inflammatory response in goat endometrial epithelial cells.

The endometrium plays an important role in the defence against invading pathogens, although the mechanisms are not clear. UFMylation is a recently discovered novel ubiquitination-like modification system that plays a pivotal role in inflammation and the immune response. The purpose of this study was to investigate the effects of UFMylation on lipopolysaccharide (LPS)-induced inflammatory responses in immortalized goat endometrial epithelial cells (gEECs). Ubiquitin-fold modifier conjugating enzyme 1 (UFM1) and DDRGK domain containing 1 (DDRGK1) were mainly localized in the luminal epithelium and glandular epithelium of mouse and goat endometrial tissues. The expression levels of UFM1, ubiquitin-like modifier activating enzyme 5 (UBA5), UFM1 specific ligase 1 (UFL1) and DDRGK1, as key components of the UFMylation system, were significantly activated by 5 µg/mL LPS-induced inflammatory response in gEECs for 6 hr. Meanwhile, the expression levels of interleukin-6 (IL-6) were significantly upregulated, and tumour necrosis factor-α (TNF-α) was significantly down-regulated after overexpression of UFM1 in gEECs. Additionally, we observed UFM1 and DDRGK1 were markedly increased on LPS-stimulated mouse endometritis in vivo. In conclusion, the current study demonstrated that UFMylation was significantly activated by LPS and might be involved in regulating inflammatory response in gEECs.

Laminin 511 Precoating Promotes the Functional Recovery of Transplanted Corneal Endothelial Cells.

Corneal endothelial dysfunction is a major cause of corneal blindness and is mainly treated by corneal transplantation. However, the global shortage of donor cornea hampers its application. Intracameral injection of cultured primary corneal endothelial cells (CECs) was recently confirmed in clinical trials. However, abnormal adhesion of the grafted CECs affects the application of this strategy. In this study, we explored if laminin 511 (LN511) improves the therapeutic function of the intracameral CEC injection for corneal endothelial dysfunction. To mimic the late stage of corneal endothelial diseases, intense scraping was developed to remove CECs and extracellular matrix of the posterior Descemet's membrane (DM) without DM removal in rabbits. Then, Dulbecco's phosphate-buffered saline (DPBS) and LN511 were intracamerally injected as the control and intervention groups, respectively. We found that the injected LN511 could settle and form a coating on the posterior surface of DM. After CEC transplantation, corneal clarity of rabbits in the LN511 group was rapidly recovered within 7 days, whereas the corneal recovery took 14 days in the DPBS group. Corneal thickness of LN511 group decreased to 413.3 ± 20.8 µm 7 days after operation, which was significantly lower than 1086.3 ± 78.6 µm of DPBS group (p < 0.01). Moreover, for the grafted CECs, LN511 promoted the rapid adhesion, tight junction formation, and expression of Na+/K+-ATPase and ZO-1. In vitro analysis revealed that the functions of LN511 on the cultured human CECs mechanistically depended on the cell density and the nuclear-cytoplasmic translocation of the Yes-associated protein. Our study demonstrated that LN511 precoating promoted the adhesion of the transplanted CECs and enhanced the functional regeneration of the corneal endothelium. Thus, our data suggested that the strategy of LN511 precoating and CECs' intracameral injection could be a potential method for the therapy of corneal endothelial dysfunction. Impact statement Intracameral injection of cultured corneal endothelial cells (CECs) is a potential alternative therapy for corneal endothelial dysfunction and has been proven to be effective in clinical trials. However, abnormal adhesion of the grafted CECs affects its application. In this study, intense scraping was developed to remove CECs and extracellular matrix of the posterior Descemet's membrane (DM) without DM removal for the therapy of late stage of corneal endothelial diseases. Laminin 511 was intracamerally injected to form a coating, improve the posterior DM, enhance the adhesion of the grafted CECs, and promote the functional regeneration of CEC transplantation through Yes-associated protein signaling.

NAD+ precursors protect corneal endothelial cells from UVB-induced apoptosis.

Excessive exposure of the eye to ultraviolet B light (UVB) leads to corneal edema and opacification because of the apoptosis of the corneal endothelium. Our previous study found that nicotinamide (NIC), the precursor of nicotinamide adenine dinucleotide (NAD), could inhibit the endothelial-mesenchymal transition and accelerate healing the wound to the corneal endothelium in the rabbit. Here we hypothesize that NIC may possess the capacity to protect the cornea from UVB-induced endothelial apoptosis. Therefore, a mouse model and a cultured cell model were used to examine the effect of NAD+ precursors, including NIC, nicotinamide mononucleotide (NMN), and NAD, on the UVB-induced apoptosis of corneal endothelial cells (CECs). The results showed that UVB irradiation caused apparent corneal edema and cell apoptosis in mice, accompanied by reduced levels of NAD+ and its key biosynthesis enzyme, nicotinamide phosphoribosyltransferase (NAMPT), in the corneal endothelium. However, the subconjunctival injection of NIC, NMN, or NAD+ effectively prevented UVB-induced tissue damage and endothelial cell apoptosis in the mouse cornea. Moreover, pretreatment using NIC, NMN, and NAD+ increased the survival rate and inhibited the apoptosis of cultured human CECs irradiated by UVB. Mechanistically, pretreatment using nicotinamide (NIC) recovered the AKT activation level and decreased the BAX/BCL-2 ratio. In addition, the capacity of NIC to protect CECs was fully reversed in the presence of the AKT inhibitor LY294002. Therefore, we conclude that NAD+ precursors can effectively prevent the apoptosis of the corneal endothelium through reactivating AKT signaling; this represents a potential therapeutic approach for preventing UVB-induced corneal damage.

Progesterone-induced RNA Hand2os1 regulates decidualization in mice uteri.

Decidualization is a critical process for successful embryo implantation and subsequent placenta formation. The characterization and physiological function of lncRNA during decidualization remain largely unknown. In the present study, we conducted RNA-sequencing analysis to compare gene expression between decidua of days 6 and 8, and normal pregnant endometrium (day 4). A total of 2332 high-confidence putative lncRNA transcripts were expressed. Functional clustering analysis of cis and trans lncRNA targets showed that differentially expressed lncRNAs may regulate multiple gene ontology terms and pathways that have important functions in decidualization. Subsequent analyses using qRT-PCR validated that eight of all lncRNAs were differentially regulated in mice uteri during decidualization, both in vivo and in vitro. Furthermore, we showed that differentially expressed lncRNA of Hand2os1 was specifically detected in stromal cells on days 2 to 5 of pregnancy and was strongly upregulated in decidual cells on days 6-8 of pregnancy. Similarly, Hand2os1 expression was also strongly expressed in decidualized cells following artificial decidualization, both in vivo and in vitro. In uterine stromal cells, P4 was able to significantly upregulate the expression of Hand2os1, but upregulation was impeded by RU486, whereas E2 appeared to have no regulating effect on Hand2os1 expression. Concurrently, Hand2os1 significantly promoted the decidual process in vitro and dramatically increased decidualization markers Prl8a2 and Prl3c1. Our results provide a valuable catalog for better understanding of the functional roles of lncRNAs in pregnant mouse uteri, as it relates to decidualization.

Ocular surface repair using decellularized porcine conjunctiva.

The primary functions of the conjunctiva embody ocular surface protection and the maintenance of the tear film equilibrium. Severe conjunctival defects such as symblepharon may impair the integrity of ocular surface and cause loss of visual functions. Here we report the use of a decellularized porcine conjunctiva (DPC) for conjunctival reconstruction in rabbit models and in clinic. Our results show that the major xenoantigens are efficiently removed, while abundant matrix components and integrated microstructures are well preserved in the DPC. These characteristics provide mechanical support and favorable histocompatibility for repairing damaged conjunctiva. The DPC application has demonstrated enhanced transplant stability and improved epithelial regeneration in severe ocular surface damage comparing to those of amniotic membrane (AM), the most frequently applied matrix for ocular surface reconstruction nowadays. In order to test the DPC performance in clinic, three patients with pterygium and one patient with symblepharon underwent transplant with DPC. The grafts in all cases were completely re-epithelized and no graft melt or fibroplasia were observed. These results suggest that the strategy we developed is feasible and effective for conjunctival reconstruction and ocular surface repair. STATEMENT OF SIGNIFICANCE: In this study, we adopted an innovative approach to prepare decellularized porcine conjunctiva (DPC). The intricate conjunctiva-specific structures and abundant matrix components were preserved in DPC, which offers favorable mechanical properties for graft. DPC has shown positive effects in ocular surface repair, which has been proven particularly in a rabbit model with severe symblepharon. Reconstructed conjunctiva by DPC exhibited epithelial heterogeneity, extremely resembling that of native conjunctiva. In addition, results from clinical studies were encouraging for pterygium and symblepharon and clinical application of DPC is promising.

CREBZF regulates testosterone production in mouse Leydig cells.

CREBZF, including the two isoforms SMILE (long isoform of CREBZF) and Zhangfei (short isoform of CREBZF), has been identified as a novel transcriptional coregulator of a variety of nuclear receptors. Our previous studies found that SMILE is expressed in the mouse uterine luminal and glandular epithelium and is upregulated by estrogen. In the present study, CREBZF was age-dependently and -specifically expressed in mouse interstitial Leydig cells during sexual maturation. The expression pattern of CREBZF exhibited an age-related increase, and SMILE was the dominant isoform in the mouse testis. Although hCG did not affect CREBZF expression, CREBZF silencing significantly inhibited hCG-stimulated testosterone production in primary Leydig cells and MLTC-1 cells. Meanwhile, the serum concentration of testosterone was significantly decreased after microinjection of lentiviral-mediated shRNA-CREBZF into the mature mouse testis. In addition, CREBZF silencing markedly decreased P450c17, 17ß-HSD, and 3ß-HSD expression following hCG stimulation in primary Leydig cells, and this inhibitory effect was obviously reversed by overexpression of CREBZF. Furthermore, CREBZF significantly upregulated the mRNA levels of Nr4a1 and Nr5a1, which are the essential orphan nuclear receptors for steroidogenic gene expression. Together our data indicate that CREBZF promotes hCG-induced testosterone production in mouse Leydig cells by affecting Nr4a1 and Nr5a1 expression levels and subsequently increasing the expression of steroidogenic genes such as 3ß-HSD, 17ß-HSD, and P450c17, suggesting a potential important role of CREBZF in testicular testosterone synthesis.

Nicotinamide inhibits corneal endothelial mesenchymal transition and accelerates wound healing.

Corneal endothelial cells (CECs) maintain the clarity of the cornea through the barrier and pump function. Ex vivo culture or injury may cause corneal endothelial-mesenchymal transition (EnMT) and lead to loss of function. In this study, we explored the effects of nicotinamide (NIC) on the wound healing of rabbit corneal endothelium and the proliferation, migration, and EnMT of cultured human CEC lines. The animal results showed that corneal clarity was rapidly recovered within seven days through topical application of NIC in the rabbits with mechanical injury of the corneal endothelium, while the control corneas remained edematous and cloudy. Whole-mounted corneal staining found the expressions of Na+/K+-ATPase, aquaporin-1, and zonula occludens-1 were mainly localized to the boundaries of regenerated endothelium in NIC-treated eyes, in contrast to the scattered staining in vehicle-treated eyes. Interestingly, we found that NIC application inhibited the expression of typical EnMT marker alpha-smooth muscle actin, which appeared in the rabbit corneal endothelial wound healing. In vitro, NIC promoted the proliferation, but not the migration, of cultured human CECs. Moreover, NIC effectively inhibited transforming growth factor beta-1-induced corneal EnMT and decreased the levels of EnMT regulators snail and slug. Therefore, our study indicates that NIC enhances corneal endothelial wound healing through the promotion of proliferation and the inhibition of EnMT, which may provide a potential pharmaceutical agent for treating corneal endothelial dysfunction.

Rapid Differentiation of Multi-Zone Ocular Cells from Human Induced Pluripotent Stem Cells and Generation of Corneal Epithelial and Endothelial Cells.

Eye is a complex organ with a highly specialized tissue structure. The establishment of human pluripotent stem cells (hPSCs) has allowed the simulation of eye development in vitro. Most differentiation works of hPSC-derived ocular cells focus on a single, tissue-specific lineage, however, that faces difficulty in reflecting the complexity of eye development. Recently, the generation of a self-formed ectodermal autonomous multi-zone of ocular cells availably mimics the process of whole-eye development. In this study, we developed a rapid defined method to induce the differentiation of multi-zone ocular cells (MZOCs) from human induced pluripotent stem cells, which specifically experienced the key progenitor stages of anterior neuroectoderm and eye field stem cells by a 2.5-dimensional culture. These differentiated cell types spanned neural retina, retinal pigment epithelium, surface ectoderm, and neural crest and lens cells. In addition, the surface ectoderm zone of MZOCs could be mechanically isolated and induced into corneal epithelial cells, and the isolated neural crest zone could be directed into corneal endothelial cells. This in vitro differentiation process vividly mimics the development of vertebrate eye, and it provides a promising model for the study of ocular morphogenesis, as well as an ideal resource of seed cells for corneal regenerative medicine.

Characteristics of New Onset Herpes Simplex Keratitis after Keratoplasty.

PURPOSE: To observe clinical characteristics and treatment outcomes of new onset herpes simplex keratitis (HSK) after keratoplasty. METHODS: Among 1,443 patients (1,443 eyes) who underwent keratoplasty (excluding cases of primary HSK) in Shandong Eye Hospital, 17 patients suffered postoperative HSK. The clinical manifestations, treatment regimens, and prognoses of the patients were evaluated. RESULTS: The incidence of new onset HSK after keratoplasty was 1.18%. Epithelial HSK occurred in 10 eyes, with dendritic epithelial infiltration in 6 eyes and map-like epithelial defects in 4 eyes. Nine eyes had lesions at the junction of the graft and recipient. Stromal necrotic and endothelial HSK occurred in 7 eyes, presenting map-shaped ulcers in the entire corneal graft and recipient (two eyes) or at the graft-recipient junction (five eyes). Confocal microscopy revealed infiltration of a large number of dendritic cells at the junction of the lesion and transparent cornea. All 10 eyes with epithelial lesions and two eyes suffering stromal lesions of ≤1/3 corneal thickness healed after systematic and local antiviral treatment. Best-corrected visual acuity and corneal graft transparency were restored. For stromal HSK with an ulcer of >1/3 corneal thickness, amniotic membrane transplantation was performed, and visual acuity and graft transparency decreased significantly. CONCLUSION: New onset HSK after keratoplasty primarily resulted in epithelial and stromal lesion, involving both the graft and recipient. Effective treatments included antiviral medications and amniotic membrane transplantation. Delayed treatment may lead to aggravated graft opacification.