In vitro synthesis of vitamin D-3 by cultured human keratinocytes and fibroblasts: action spectrum and effect of AY-9944

With delineation of the photochemical events occurring in the skin after ultraviolet exposure, there has been increased interest in the skin's role in the vitamin D-3-endocrine system. We provide here in vitro conditions for the generation of both labelled (from [3H]acetate) and unlabelled vitamin D-3 in cultures of human keratinocytes and fibroblasts. Sterol precursors and photoproducts in irradiated and non-irradiated cultures are identified by co-chromatography, ultraviolet absorbance spectra, thermal conversion characteristics of previtamin D-3 and mass spectrometry. Because the conversion of 7-dehydrocholesterol to cholesterol is more efficient in vitro than in vivo, the specific delta 7 inhibitor, AY-9944, was added in non-toxic doses to modulate 7-dehydrocholesterol content. Both cell types were equally capable of generating photoproducts, depending on the amount of 7-dehydrocholesterol present. The 290 +/- 5 and 295 nm filters were much more efficient than the 305 nm filter for generating previtamin D-3 and vitamin D-3 in fibroblasts. In contrast, the 305 nm filter was as efficient as the 290 +/- 5 and 295 nm filters in keratinocytes, where it yielded previtamin D-3, with much less lumisterol and tachysterol than appeared with the shorter-wavelength filters. The amount of lumisterol and tachysterol versus previtamin D-3 formed in both cell types was dependent on the total energy applied, with lower energies (less then 1 J/cm2) favoring previtamin D-3 over the other photoproducts. The use of cultured cells provides a system whereby the regulation of vitamin D-3 synthesis by extracutaneous factors can be studied in a homogeneous setting.     

A preliminary note on the use of milk substitutes in the early weaning of dromedary camels

The experiment was performed using two young male camels which weighed 24 and 36 kg respectively at birth. Each young camel was weighed and abruptly separated from its mother after 30 days of nursing (or at 1 month of age). The weaned calves were fed milk substitutes prepared commercially for lambs by Mabarot Chemical and Veterinary Products (Israel). The weanling camels averaged daily weight gains of 0.400 and 1.0 kg per day respectively during the 30 day initial period when the milk substitutes were used. Following the period when milk substitutes were used, the camels achieved normal growth to arrive at 135 and 145 kg respectively at 6 months of age.     

Minireview: Metabolic control of the reproductive physiology: Insights from genetic mouse models

This article is part of a Special Issue Energy Balance.     

Processive replication of single-stranded DNA templates by the herpes simplex virus-induced DNA polymerase

The DNA polymerase encoded by herpes simplex virus 1 consists of a single polypeptide of Mr 136,000 that has both DNA polymerase and 3'----5' exonuclease activities; it lacks a 5'----3' exonuclease. The herpes polymerase is exceptionally slow in extending a synthetic DNA primer annealed to circular single-stranded DNA (turnover number approximately 0.25 nucleotide). Nevertheless, it is highly processive because of its extremely tight binding to a primer terminus (Kd less than 1 nM). The single-stranded DNA-binding protein from Escherichia coli greatly stimulates the rate (turnover number approximately 4.5 nucleotides) by facilitating the efficient binding to and extension of the DNA primers. Synchronous replication by the polymerase of primed single-stranded DNA circles coated with the single-stranded DNA-binding protein proceeds to the last nucleotide of available 5.4-kilobase template without dissociation, despite the 20-30 min required to replicate the circle. Upon completion of synthesis, the polymerase is slow in cycling to other primed single-stranded DNA circles. ATP (or dATP) is not required to initiate or sustain highly processive synthesis. The 3'----5' exonuclease associated with the herpes DNA polymerase binds a 3' terminus tightly (Km less than 50 nM) and is as sensitive as the polymerase activity to inhibition by phosphonoacetic acid (Ki approximately 4 microM), suggesting close communication between the polymerase and exonuclease sites.     

Nerve Growth Factor Effect on Adenosine Transport in Cultured Chromaffin Cells

Chromaffin cells both recently isolated or in culture present a high-affinity adenosine transporter with a Km value of 1 microM. When cells were exposed to nerve growth factor (NGF; 10 ng/ml), the adenosine transporter affinity decreased to 3 microM. This value was maintained from 3 days after plating to the end of the culture period. A change in the transport capacity was observed, with a significant increase (approximately 200-260%) in NGF-cultured cells throughout the period studied.     

Modelling mass transfer and agitator performance in multiturbine fermentors

A methodology for mathematically analyzing agitator performance and mass transfer in large multiturbine production fermentors is presented. The application of this approach provides a method for determining axial dissolved oxygen profiles under conditions of known mass transfer rates as a function of agitation-aeration characteristics. A stagewise approach is used which divides the fermentor into a series of mixing cells. This allows for each turbine and mixing cell to be individually optimized. The model also permits the determination of the mass transfer coefficient for each turbine based upon limited dissolved oxygen data. The primary limitation of this approach rests in the limited data and correlations available for multiturbine systems. The structure of the modelling approach can serve as a basis for testing single turbine correlations and adapting them to multiturbine systems. The step-by-step details of the mathematical analysis are presented and interpreted. A series of computer simulations demonstrate the effect of typical fermentor operating variables on the axial dissolved oxygen profile. Further simulations demonstrate the effect of modifying agitator blade numbers on the dissolved oxygen profile and agitator power requirement.