Ultrasound thermal damage to rabbit corneas after simulated phacoemulsification

PURPOSE: To determine rabbit cornea thermal tolerance and evaluate the effects of ultrasound (US) on this tissue after applying defined US heat doses. SETTING: Eye Clinic; Anatomy Histology and Forensic Medicine, University of Florence, Florence, Italy. METHODS: Hyperthermia was induced in rabbit corneas using US, simulating a phacoemulsification procedure. The US power was set at 100% in continuous mode, and temperature values were reached within 10 seconds of the onset of US treatment. Corneal surface temperatures were continuously monitored and recorded by thermographic registration. The eyes of 16 rabbits were examined: 4 controls, 8 treated at 40 degrees C for 10 seconds, 8 treated at 50 degrees C for 10 seconds, and 12 treated at 60 degrees C for 10 seconds. All 32 corneal buttons were removed and prepared for light microscopic evaluation with hematoxylin and eosin staining, trichromic staining, and zinc iodide-osmium tetroxide solution. The 12 corneas treated at 60 degrees C for 10 seconds were also processed for immunohistochemical analysis. RESULTS: Corneas at 40 degrees C for 10 seconds were grossly and histologically normal and were not different from control corneas. Corneas at 50 degrees C for 10 seconds showed initial stromal damage with collagen disorganization, mild stromal edema, and initial signs of keratocyte damage. Half of the corneas at 60 degrees C for 10 seconds were examined at time 0 and the other half after 1 week. At time 0, massive corneal damage with epithelial cell edema, collagen disorganization, severe stromal edema, intrastromal vacuole formation, plump keratocyte nuclei, and endothelial cell detachment were found, as was a severely impaired nerve plexus. At 1-week follow-up, corneas showed persistent stromal and endothelial cell edema with an increase activated keratocytes and mitotic features in the stroma and the epithelial layer. CONCLUSIONS: Rabbit corneas showed a considerable tolerance to US damage up to 50 degrees C. Higher thermal doses produced severe histological damage, even though corneas showed a considerable plasticity due to their regenerative capacity.     

BXL-628, a vitamin D receptor agonist effective in benign prostatic hyperplasia treatment, prevents RhoA activation and inhibits RhoA/Rho kinase signaling in rat and human bladder

BACKGROUND: BXL-628 is a calcitriol analog shown to decrease prostate growth in preclinical and clinical studies. BPH symptoms are generated not only by prostate overgrowth but also by bladder overactivity, resulting from an increased RhoA/Rho-kinase signaling. Because bladder smooth muscle cells express VDR, we studied effects of BXL-628 on this pathway. METHODS: RhoA and Rho-kinase gene expression and functional activity were studied in rat and human bladder smooth muscle by real-time RT-PCR, immuno-kinase assays, western blot analysis, confocal microscopy, in vitro contractility, and cell migration. RESULTS: In bladder smooth muscle, carbachol responsiveness was delayed and Rho-kinase activity reduced by BXL-628 treatment because of impaired RhoA membrane translocation and activation. Accordingly, RhoA-mediated biological functions, such as cell migration and cytoskeleton remodeling were also inhibited by BXL-628. CONCLUSIONS: BXL-628 inhibits RhoA/Rho-kinase signaling, a calcium sensitizing pathway, suggesting its possible clinical use in the treatment of altered bladder contractility often associated with BPH-induced lower urinary tract symptoms.     

Guidelines for reporting experiments involving animals: the ARRIVE guidelines

British Journal of Pharmacology (BJP) is pleased to publish a new set of guidelines for reporting research involving animals, simultaneously with several other journals; the ‘ARRIVE’ guidelines (Animals in Research: Reporting In Vivo Experiments). This editorial summarizes the background to the guidelines, gives our view of their significance, considers aspects of specific relevance to pharmacology, re-states BJP''s guidelines for authors on animal experiments and indicates our commitment to carrying on discussion of this important topic. We also invite feedback via the British Pharmacological Society website.     

Role of the signal peptide in the synthesis and processing of the glucagon-like peptide-1 receptor

Background and purpose:

The glucagon-like peptide-1 receptor (GLP-1R) belongs to Family B of the G protein-coupled receptor superfamily and is a target for treatment of type 2 diabetes. Family B G protein-coupled receptors contain a putative N-terminal signal peptide, but its role in receptor synthesis and trafficking are unclear. Further, the signal peptide is not cleaved in at least one family member.

Experimental approach:

We examined receptor glycosylation and the role of the signal peptide in GLP-1R synthesis and trafficking using constructs containing epitope tags at the N- and/or C-terminus and in which the signal peptide sequence was either present or absent.

Key results:

The signal peptide was absolutely required for GLP-1R synthesis but could be substituted to some extent by increasing positive charge in the N-terminal region of the receptor flanking the signal peptide. The signal peptide is cleaved during synthesis and processing of the receptor. An enhanced GFP-epitope tag at the N-terminus of the receptor permitted synthesis of the receptor but blocked signal peptide cleavage and prevented trafficking to the plasma membrane. Cleavage site mutation allowed synthesis of a full-length receptor, blocked signal peptide cleavage and caused retention within the endoplasmic reticulum.

Conclusions and implications:

Signal peptide cleavage was not essential for receptor synthesis but was obligatory for processing and trafficking of receptors to the plasma membrane. Further, the GLP-1R is subject to N-linked glycosylation and only the mature, fully glycosylated form of the receptor is present in the plasma membrane. Inhibition of glycosylation prevents processing and cell surface expression of the GLP-1R.     

Relationship between the neuroprotective effects of insulin-like growth factor-1 and 17β-oestradiol in human neuroblasts

Insulin-like growth factor-1 (IGF-1) and oestrogens interact with each other as neuroprotective factors. We have previously demonstrated that 17β-oestradiol protects against β-amyloid and oxidative stress toxicity and increases the amount of cell cholesterol in human foetal neuroblasts (FNC). The present study aimed: (i) to assess the protective effects of IGF-1 in FNC cells; (ii) to investigate the relationship between IGF-1 and 17β-oestradiol; and (iii) to determine whether cholesterol was a major mediator of the effects of IGF-1, similarly to 17β-oestradiol. We found that IGF-1 effectively exerts neuroprotective effects in FNC cells. We also demonstrated that the IGF-1 receptor (IGF-1R) pathway is needed to maintain oestrogen-mediated neuroprotection. Finally, we found that, opposite to 17β-oestradiol, IGF-1 did not cause a significant increase in cell cholesterol. These findings indicate that a cross-talk between IGF-1 and 17β-oestradiol occurs in FNC cells. In particular, the activation of the IGF-1R cascade appears to be fundamental to warrant 17β-oestradiol-mediated neuroprotection, even though cell cholesterol does not play a major role as an effector of this pathway.     

Cell-based therapy in Alzheimer's disease: can human fetal cholinergic neurons “untangle the skein”?

正Alzheimer’s disease(AD)is a devastating neurodegenerative disorder and the most common form of old-age dementia.The disease is characterized by a progressive decline in cognitive functions,gradual loss of memory and ability to perform everyday activities,and leads to inevi-     

Giant condyloma acuminatum of the anorectum: successful radical surgery with anal reconstruction

Buschke-L?wenstein tumor, or giant condyloma acuminatum, is a relatively uncommon lesion of the anus with aggressive local invasive behavior which may present as a large warty tumor of the genital region with expansive and destructive growth. Many sporadic reports have been published suggesting various therapeutic strategies. We report a case of Buschke-L?wenstein tumor treated with conservative surgery followed by reconstructive procedures without a loop colostomy     

Metastasis of sigmoid colon cancer in cryptorchid testis: report of a case

Isolated testicular metastasis from colorectal cancer is considered an unusual event. In this case report we describe for the first time a metastasis from an adenocarcinoma of the sigmoid colon to a cryptorchid testis. The patient developed a painless testicular nodule three years after the diagnosis of primary sigmoid colon cancer. Recent reports have suggested that the incidence of genitourinary abnormalities in human males has increased over the past 50 years; in particular, cryptorchid testes increase the clinical risk factors for primary or metastatic testicular cancer. In conclusion, there should be awareness of the risk of metastasis of colorectal cancer to the testis in the workup of patients with testicular symptoms. Furthermore, patients with colorectal cancer and cryptorchidism should be managed with a single surgical intervention: when the primary colorectal tumor is removed, the cryptorchid testicle should also be removed to reduce the risk of late metastases.     

Effects of riboflavin/UVA corneal cross‐linking on keratocytes and collagen fibres in human cornea

Purpose: To evaluate the effects of corneal cross‐linking on keratocytes and collagen fibres in human corneas. Methods: Fifteen corneal buttons were examined. Ten were from patients with keratoconus submitted to penetrating keratoplasty and five of them were treated with cross‐linking 6 months before penetrating keratoplasty. Five normal corneal buttons from healthy donors were used as controls. All samples were prepared for TUNEL assay and Western blot analysis for the detection of keratocyte apoptosis and immunohistochemical analysis for the morphological evaluation of keratocytes and collagen fibre diameter. Results: Normal corneas exhibited no TUNEL‐positive keratocytes and keratoconic and cross‐linked corneas showed moderate apoptotic cells mainly in the anterior part of the stroma. This apoptotic trend was confirmed by the cleavage of poly (ADP‐ribose) polymerase assessed using Western blot. The Ki‐67 staining showed a significant increase in the keratocyte proliferation in cross‐linked corneas compared with normal and keratoconus. In cross‐linked corneas CD34‐positive keratocytes were regularly distributed throughout the whole corneal stroma as in the control, and keratoconus was associated with patchy loss of immunoreactivity. The immunohistochemical analysis of collagen type I showed a significant increase in fibre diameter of cross‐linked corneas compared with control and keratoconus. Conclusion: Corneal cross‐linking leads to keratocyte damage; after 6 months a repopulation by proliferating cells, a distribution of CD34‐positive keratocytes as in control and an increase in collagen fibre diameter were observed. These modifications are the morphological correlate of the process leading to an increase in biomechanical stability.