Clive R. Hamlin

Apparent Accelerated Aging of Human Collagen in Diabetes Mellitus

The chronologic ages of human subjects were determined experimentally by enzymatic digestion of tendon collagen samples. Determined age closely matched actual age for individuals dying with a variety of major diseases. Juvenile diabetics did not fit this pattern; their experimentally determined ages were significantly greater than their actual ages. This raises the possibility of relationships between diabetes mellitus, changes in connective tissue, and accelerated aging.

Estrogen bows to a new master : The role of gonadotropins in alzheimer pathogenesis

Abstract: Epidemiological data showing a predisposition of women to develop Alzheimer disease (AD) led many researchers to investigate the role of sex steroids, namely estrogen, in disease pathogenesis. Although there is circumstantial support for the role of estrogen, the unexpected results of the Womens Health Initiative (WHI) Memory Study, which reported an increase in the risk for probable dementia and impaired cognitive performance in postmenopausal women treated with a combination of estrogen and progestin, have raised serious questions regarding the protective effects of estrogen. Although explanations for these surprising results vary greatly, the WHI Memory Study cannot be correctly interpreted without a complete investigation of the effects of the other hormones of the hypothalamic‐pituitary‐gonadal (HPG) axis on the aging brain. Certain hormones of the HPG axis, namely, the gonadotropins (luteinizing hormone and follicle‐stimulating hormone), are not only involved in regulating reproductive function via a complex feedback loop but are also known to cross the blood‐brain barrier. We propose that the increase in gonadotropin concentrations, and not the decrease in steroid hormone (e.g., estrogen) production following menopause/andropause, is a potentially primary causative factor for the development of AD. In this review, we examine how the gonadotropins may play a central and determining role in modulating the susceptibility to, and progression of, AD. On this basis, we suggest that the results of the WHI Memory Study are not only predictable but also avoidable by therapeutically targeting the gonadotropins instead of the sex steroids.

Diagnostic and prognostic value of human prion detection in cerebrospinal fluid

Several prion amplification systems have been proposed for detection of prions in cerebrospinal fluid (CSF), most recently, the measurements of prion seeding activity with second‐generation real‐time quaking‐induced conversion (RT‐QuIC). The objective of this study was to investigate the diagnostic performance of the RT‐QuIC prion test in the broad phenotypic spectrum of prion diseases.

Determination of human chronological age by study of a collagen sample

Abstract The stabilization (‘cross-linking’) of insoluble human collagen has been shown to progress so predictably that it can form the basis of a method to determine an individuals age by study of a tissue specimen.

The effect of hydrocortisone acetate loaded poly(DL-lactide) films on the inflammatory response☆

Abstract Poly ( DL -lactide) (DL-PL) films loaded with hydrocortisone acetate (HCA) were prepared and the effects of this drug releasing system on the inflammatory and healing responses were investigated in vivo using a cage implant technique. Cytochemical staining procedures and extracellular enzyme activity assays were performed in order to characterize the cellular events in the inflammatory exudate over a 21-day implantation period. The results showed that the HCA/DL-PL sustained drug releasing system dramatically inhibited all aspects of inflammation. The white cell concentration in the HCA/DL-PL exudates was markedly lower than the white cell concentrations of both the empty cage and DL-PL controls throughout the implantation period. The drug inhibited the accumulation of all types of leukocytes at the site of inflammation including polymorphonuclear leukocytes, macrophages and lymphocytes. Additionally, examination of the retrieved cages 21 days after implantation revealed minimal connective tissue development around the HCA/DL-PL implants compared to the two sets of controls. This indicated that the healing phase of inflammation was being repressed.

Partial characterization of the age-related stabilizing factor of post-mature human collagen-I. by the use of bacterial collagenase

Abstract A stabilizing factor that causes resistance to digestion becomes increasingly important in collagen of post-mature individuals as they age. This is demonstrated with paired diaphragm tendon and the dura mater from single individuals. The factor is rapidly and irreversibly lost when purified collagen is heated to70°C. Contact with 70% formic acid quickly abolished any detectable age differences, yet strong acids and bases did not disrupt the stabilizing factor. Treatment with cyanogen bromide in 70% formic acid at 30°C failed to solubilize the collagen even when samples were first heated to 100°C in the presence of 0·01 M sodium hydroxide. Other treatments, including exposure to sodium borohydride or glycosidases, had no detectable effect on teh stabilizing factor. Increasing calcium concentration enhanced the rate of enzymatic digestion when using bacterial collagenase. Below 0·5 M calcium, age differences are readily observed with collagenase, but the differences are lost at higher concentrations. The age differences are regained when the calcium ion concentration is reduced. This rapid reversal in contrast to the irreversible loss of the age difference caused by 70% formic acid treatment or collagen denaturants. These two latter treatments may not abolish the stabilization factor but could instead modify the collagen structure to the point where it can no longer retard the rate of collagenase digestion.

Aging of collagen: comparative rates in four mammalian species.

Abstract Resistance to collagenase digestion was used to compare rates of collagen aging in rat, dog, macaque, and man. The apparent rates were different for each species. Rat collagen was uniformly and readily digested, even at low calcium ion concentration where human collagen is digested slowly. Dog, macaque and human collagens all showed evidence of a progressive, age-related, increased stabilization. Some older dog specimens showed a much greater extent of stabilization than human specimens of the same chronological age, but similar if relative lifespans are used. These results demonstrate the problems caused by uncritically extrapolating an age-related observation from a short-lived species such as rat to a long-lived species such as human.

Human collagen digestion: I. Nature of the digestion kinetics as a function of age and structure

The digestion of human collagen by bacterial collagenase was studied in order to relate collagen structure to the digestion kinetics. The digestion was shown to be first order in collagen and in collagenase on given material from run to run. There was no product inhibition and no enzyme deactivation. However, the rate changed with age, and also within a run as a function of conversion. It was shown that the normalized time (time X enzyme concentration), to reach a given amount of digestion can be plotted as a linear function of the sample age, from 20 to 70 years. The digestion broke down into two regions. The first was a rapid first order region which amounted to about 10% of the collagen. The second region could be graphed as a second order plot to obtain a straight line. This implied that the rate of digestion was functionally second order in collagen; first order in concentration and first order in conversion of collagen.

A rare case of rapidly progressive dementia with elevated RT-QuIC and negative 14-3-3 and tau proteins

ABSTRACT Creutzfeldt-Jakob disease (CJD) is characterized by rapidly progressing dementia with death usually occurring within 6 months. There is no verified disease-specific pre-mortem diagnostic test besides brain biopsy. We describe a 66 y old previously high functioning male who presented with a 5 month history of rapidly progressive dementia. Neurological examination revealed a score of 19/30 on MOCA testing. An extensive workup into various causes of dementia including electroencephalography and imaging studies was unremarkable. The cerebrospinal fluid was sent to National Prion Disease Center and it revealed elevated RT-QuIC levels with negative 14-3-3 and T tau proteins. Based on literature review, our case is one of few living subjects with elevated RT-QuIC levels and negative 14-3-3 and tau proteins.

Human collagen digestion: II. Theoretical model and application to data

A kinetic mechanism for the digestion of collagen by bacterial collagenase has been proposed which is consistent with experimental data. Arguments are presented justifying the application of a homogeneous approach to the digestion of insoluble collagen. The mechanism takes account of increased kinetic blockages with aging and enzyme complexity. It is shown to be applicable to collagen from various tissues, including relatively impure preparations. However, the mechanism is not applicable to collagen from the immature period of the lifespan. Kinetic blockages are shown to increase three- to four-fold from age 20 to 70 years, within the main helical region of the collagen molecule. There is no increase of kinetic blockages in the N-terminal region of the molecule over this age range.