High-performance size-exclusion chromatography of peptides

Gel filtration on soft gels has been employed for over 40 years for the separation, desalting and molecular weight estimation of peptides and proteins. Technical improvements have given rise to high-performance size-exclusion chromatography (HPSEC) on rigid supports, giving more rapid run times and increased resolution. Initially, these packings were more suitable for the separation of proteins than of peptides, but supports that operate in the fractionation range

Comparative effects of GLP-1 and GIP on cAMP production, insulin secretion, and in vivo antidiabetic actions following substitution of Ala8/Ala2 with 2-aminobutyric acid

The two major incretin hormones, glucagon-like peptide-1 (GLP-1), and glucose-dependent insulinotropic polypeptide (GIP), are currently being considered as prospective drug candidates for treatment of type 2 diabetes. Interest in these gut hormones was initially spurred by their potent insulinotropic activities, but a number of other antihyperglycaemic actions are now established. One of the foremost barriers in progressing GLP-1 and GIP to the clinic concerns their rapid degradation and inactivation by the ubiquitous enzyme, dipeptidyl peptidase IV (DPP IV). Here, we compare the DPP IV resistance and biological properties of Abu(8)/ Abu(2) (2-aminobutyric acid) substituted analogues of GLP-1 and GIP engineered to impart DPP IV resistance. Whereas (Abu(8))GLP-1 was completely stable to human plasma (half-life > 12h), GLP-1, GIP, and (Abu(2))GIP were rapidly degraded (half-lives: 6.2, 6.0, and 7.1 h, respectively). Native GIP, GLP-1, and particularly (Abu(8))GLP-1 elicited significant adenylate cyclase and insulinotropic activity, while (Abu(2))GIP was less effective. Similarly, in obese diabetic (ob/ob) mice, GIP, GLP-1, and (Abu(8))GLP-1 displayed substantial glucose-lowering and insulin -releasing activities, whereas (Abu(2))GIP was only weakly active. These studies illustrate divergent effects of penultimate amino acid Ala(8)/Ala(2) substitution with Abu on the biological properties of GLP-1 and GIP, suggesting that (Abu(8))GLP-1 represents a potential candidate for future therapeutic development. (C) 2004 Elsevier Inc. All rights reserved.

An investigation into the role of Bcl-2 in neuroendocrine differentiation

INTRODUCTION: In addition to its role in apoptosis suppression, Bcl-2 has been reported to be co-expressed with neuroendocrine markers in several tissues, leading to speculation that this oncoprotein may promote neuroendocrine differentiation. AIM: This study investigated whether Bcl-2 modulated neuroendocrine biopeptide expression. METHODS: Levels of chromogranin A, neurone specific enolase, protein gene peptide 9.5, pancreatic polypeptide, and the chromogranin-derived peptides, intervening peptide and vasostatin-1 were examined by immunocytochemistry in rat phaeochromocytoma (PC12) cell lines genetically engineered to over-express Bcl-2 and their mock-transfected controls. Intensity of fluorescence was graded using a semi-quantitative scale from (-) indicating negative expression to (+++) indicating intense positivity. RESULTS: Mann-Whitney U analysis indicated that no significant differences in expression existed between control and Bcl2 over-expressing cell lines for any of the six peptides examined. CONCLUSIONS: The results of this study do not support the hypothesis that Bcl-2 promotes the acquisition of a neuroendocrine phenotype.

REporting and calibration of post-bomb C-14 data

The definitive paper by Stuiver and Polach (1977) established the conventions for reporting of 14C data for chronological and geophysical studies based on the radioactive decay of 14C in the sample since the year of sample death or formation. Several ways of reporting 14C activity levels relative to a standard were also established, but no specific instructions were given for reporting nuclear weapons testing (post-bomb) 14C levels in samples. Because the use of post-bomb 14C is becoming more prevalent in forensics, biology, and geosciences, a convention needs to be adopted. We advocate the use of fraction modern with a new symbol F14C to prevent confusion with the previously used Fm, which may or may not have been fractionation corrected. We also discuss the calibration of post-bomb 14C samples and the available datasets and compilations, but do not give a recommendation for a particular dataset.

Expression of the VEGF gene family during retinal vaso-obliteration and hypoxia.

Vascular insufficiency and retinal ischaemia precede many proliferative retinopathies and stimulate secretion of vasoactive growth factors. Vascular endothelial growth factor (VEGF) plays a major role and we therefore investigated the other members of the VEGF family: Placental growth factor (PlGF), VEGF-B, -C, and -D, and platelet derived growth factors (PDGF) A and B. Neonatal mice were exposed to hyperoxia for 5 days and then returned to room air (resulting in acute retinal ischaemia). RT-PCR demonstrated that all the members of the VEGF family are expressed in the retina and in situ hybridization (ISH) located their mRNAs primarily in ganglion cells. Similarly to VEGF itself, VEGF-C, PDGF-A, and PDGF-B were upregulated during retinal ischaemia (P < 0.05). Only PlGF gene expression increased during hyperoxia (P < 0.01). The expression pattern of these growth factors suggests a role in the normal retina and during vaso-obliterative and ischaemic phases.

Conditioning film and environmental effects on the adherence of Candida spp. to silicone and poly(vinylchloride) biomaterials.

The reported incidence of colonization of oropharyngeal medical devices with Candida spp. has increased in recent years, although few studies that have systematically examined the adherence of yeast cells to such biomaterials, the primary step in the process of colonization. This study, therefore, examined the effects of oropharyngeal atmospheric conditions (5% v/v carbon dioxide) and the presence of a salivary conditioning film on both the surface properties and adherence of Candida albicans, Candida krusei and Candida tropicalis to PVC and silicone. Furthermore, the effects of the salivary conditioning film on the surface properties of these biomaterials are reported. Growth of the three Candida spp. in an atmosphere containing 5% v/v CO2 significantly increased their cell surface hydrophobicity and reduced the zeta potential of C. albicans and C. krusei yet increased the zeta potential of C. tropicalis (p < 0.05). Furthermore, growth in 5% v/v CO2 decreased the adherence of C. tropicalis and C. albicans to both PVC and silicone, however, increased adherence of C. krusei (p < 0.05). Pre-treatment of the microorganisms with pooled human saliva significantly decreased their cell surface hydrophobicity and increased their adherence to either biomaterial in comparison to yeast cells that had been pre-treated with PBS (p < 0.05). Saliva treatment of the microorganisms had no consistent effect on microbial zeta potential. Interestingly, adherence of the three, saliva-treated Candida spp. to saliva-treated silicone and PVC was significantly lower than whenever the microorganisms and biomaterials had been treated with PBS (p < 0.05). Treatment of silicone and PVC with saliva significantly altered the surface properties, notably reducing both the advancing and receding contact angles and, additionally, the microrugosity. These effects may contribute to the decreased adherence of saliva-treated microorganisms to these biomaterials. In conclusion, this study has demonstrated the effects of physiological conditions within the oral cavity on the adherence of selected Candida spp. to biomaterials employed as oropharyngeal medical devices. In particular, this study has ominously shown that these materials act as substrates for yeast colonization, highlighting the need for advancements in biomaterial design. Furthermore, it is important that physiological conditions should be employed whenever biocompatibility of oropharyngeal biomaterials is under investigation. © 2001 Kluwer Academic Publishers.

The intraocular lens: challenges in the prevention and therapy of infectious endophthalmitis and posterior capsular opacification

Cataract is the leading cause of visual impairment worldwide. In the UK, some 30% of the population over 65 years of age have visually impairing cataract. Importantly, 88% of those with treatable visual impairment from cataract are not in contact with any ocular healthcare service, representing a major potential healthcare need [1]. In the USA, it has been estimated that 17.2% of the population (approximately 20.5 million) over 40 years of age have cataract in either eye and by 2020, this number is expected to rise to 30.1 million. Currently, cataract is responsible for 60% of Medicare costs associated with vision [2]. Furthermore, as the populations of industrialized countries such as the UK and the USA continue to age, the costs associated with treatment of cataract can only be expected to increase. Consequently, the development of the intraocular lens to replace the cataractous lens and the advances in intraocular lens design and implantation represent a major development in cataract treatment. However, despite such advances, cataract surgery is not without complications, such as postoperative infectious endophthalmitis, a rare but potentially devastating condition, and posterior capsular opacification, a less serious but much more common problem. This review will examine the epidemiology of cataracts, the polymeric construction of intraocular lenses implanted during cataract surgery and the complications of postoperative infectious endophthalmitis and posterior capsular opacification with regard to therapeutic interventions and prophylactic strategies. Advances in biomaterial design and function will be discussed as novel approaches to prevent such postoperative complications.