1,25-dihydroxyvitamin D3 (1,25(OH)2D3) is a potent inhibitor of keratinocyte proliferation, as well as a stimulator of epidermal terminal differentiation. In the present studies, we investigated the influence of epidermal growth factor (EGF) and insulin on the antiproliferative and differentiation activities of 1,25(OH)2D3. Our results indicate the following: (1) EGF caused a dramatic potentiation of the 1,25(OH)2 D3-induced inhibition of 3H-thymidine incorporation into keratinocytes in a dose-dependent manner; (2) insulin acted antagonistically on the EGF-dependent potentiation of the 1,25(OH)2D3-induced antiproliferative activity; (3) transforming growth factor-alpha potentiated 1,25(OH)2D3-induced antiproliferative activity similar to EGF; (4) the EGF effect was not dependent upon 1,25(OH)2D3 receptor mRNA up-regulation; and (5) removal of insulin from medium supplemented with growth factors significantly potentiated the 1,25(OH)2D3-induced inhibition on the number of basal cells and the 1,25(OH)2D3-dependent cornified envelope formation. In conclusion, the antiproliferative activity of 1,25(OH)2D3 in cultured normal human keratinocytes is greatly enhanced by EGF or transforming growth factor-alpha and reduced by insulin. Insulin also inhibits 1,25(OH)2D3-induced terminal differentiation of cultured normal human keratinocytes.

We have used a clonal intestmal epithehal cell hne (IEC-18) to study the mechanism of action of 1,25_dihydroxyvitamm D, (1,25(OH)zD) in vitro. 1 ,25(OH)ZD (IO-' M) elevated by over IO-fold the mRNA levels for the cytochrome P450 component (P45Occ24) of the 1 ,25(OH)zD-24-hydroxylase. Increased P45Occ24 mRNA levels were detectable at 6 h and peaked at 36 h. Below a concentration of IO-' M, 1,25(OH),D had almost no effect. However, additton of phorbol ester for 2 h made the intestine responsive to 1,25(OH)ZD concentrations as low as lo-' M.

Psoriasis is a skin disorder characterized by hyperproliferation of epidermal keratinocytes. 1 alpha,25-dihydroxyvitamin D3 (1 alpha,25(OH)2D3) and its analogs have been shown to inhibit keratinocyte proliferation in vitro and to be therapeutically effective for the treatment of psoriasis. Some patients with psoriasis, however, do not have a favorable response to 1 alpha,25 (OH)2D3 therapy. To evaluate the differential responsiveness to 1 alpha (OH)2D3 treatment, we examined the expression of vitamin D receptor mRNA in psoriatic lesions by reverse transcription-polymerase chain reaction using glyceraldehyde-3-phosphate dehydrogenase as an internal control. In this double-blind clinical trial, we recruited 18 patients who received topical treatment of 1 alpha,25(OH)2D3 (15 microgram/g Vaseline) or placebo on separated psoriatic lesions for 8 weeks. In patients who showed >90% clinical improvements of their psoriatic lesions with 1 alpha,25(OH)2D3 (n=9), an increase of 130+/-37% in vitamin D receptor mRNA level was observed in 1 alpha,25(OH)2D3-treated lesions when compared with the corresponding placebo controls. There was no increase in vitamin D receptor mRNA level in the lesions treated with this drug in patients who did not respond to the treatment. These data suggest that the antiproliferative activity of 1 alpha,25(OH)2D3 is closely associated with the expression of its cognate receptor.

It is well documented that 1 alpha,25-dihydroxyvitamin D3 (1 alpha,25[OH]2D3), the most active vitamin D metabolite, inhibits epidermal keratinocyte proliferation and promotes differentiation. 1 alpha,25(OH)2D3 can be produced in keratinocytes from 25-hydroxyvitamin D3 by the enzyme 25-hydroxyvitamin D3-1 alpha-hydroxylase (1-OHase). Hydroxylation of 1 alpha,25(OH)2D3 by 25-hydroxyvitamin D3-24-hydroxylase (24-OHase), the first step in the catabolic pathway of 1 alpha,25(OH)2D3 could significantly reduce the intracellular concentration of 1 alpha,25(OH)2D3. Therefore, the expression of 24-OHase could have a critical regulatory role in 1 alpha,25(OH)2D3-dependent gene expression. As a first step to examine this possibility, the steady state level of 24-OHase mRNA in cultured human keratinocytes (CHK) was investigated. 24-OHase mRNA was not detected in control CHK. 1 alpha,25(OH)2D3 caused a dose- and time-dependent increase in 24-OHase mRNA level. The highest accumulation of 24-OHase mRNA was observed in CHK treated with 0.1-1 microM 1 alpha,25(OH)2D3. The level of 24-OHase mRNA reached a plateau 12-24 hr after 1 alpha,25(OH)2D3 treatment. 1 beta,25-dihydroxyvitamin D3, the stereoisomer of 1 alpha,25(OH)2D3, failed to induce 24-OHase mRNA expression significantly. In addition to 24-OHase mRNA, a 1.0-kb mRNA hybridized strongly with both rat and human 24-OHase cDNA probes. The origin of this 1.0-kb message is unknown at present, however, it was regulated by 1 alpha,25(OH)2D3. These results demonstrate that 1 alpha,25(OH)2D3 up-regulates the expression of 24-OHase mRNA, and this may be an important first step in the initiation of catabolism of 1 alpha,25(OH)2D3 in human keratinocytes.

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Three-dimensional computed tomography has been used in both preoperative planning of mandibular distraction osteogenesis and in the evaluation of postoperative resolution of tongue-based airway obstruction. The authors present a case report using software to predict postdistraction airway volume during virtual surgical planning (VSP) of mandibular distraction osteogenesis in a 7 year old. The predicted increase in airway volume derived from VSP was 33.57% (1716 mm(3) preoperatively to 2292 mm(3) postvirtual distraction). Based on the three-dimensional computed tomography, the actual airway volume increased to 2211 mm(3) postoperatively, a 28.85% increase.The implications of this advancing technology are far-reaching. An illustrative case is presented herein to demonstrate the efficacy of the airway prediction and its limitations. The authors believe that, with continued investigation, this novel approach may be a standard feature of all VSP sessions for the treatment of tongue-based airway obstruction.

The transport of phosphate by primary cultures of renal cells from young (5-6 weeks) and adult (10-12 months) rats was studied. Renal tubule cells isolated from young and adult groups exhibited typical epithelial morphology and similar growth rates. The Na-dependent phosphate uptake was saturable with a Km of 5-7 microM over a substrate range of 1-500 microM. A decrease in Na-dependent phosphate uptake in adult cells (30%) was found compared to that of young cells. The Na-independent component of phosphate uptake did not vary with age. In addition, the inhibition of phosphate uptake by a variety of compounds (ouabain, gramicidin, 2,4-dinitrophenol, KCN, and arsenate) were similar in both age groups. Kinetic analysis showed that a significant reduction in Vmax (4.4 +/- 0.4 vs. 3.1 +/- 0.2 nmol Pi/mg protein/10 min in young and adult cells, respectively), but not Km, resulted in this decreased uptake of phosphate in adult groups. There was no difference in the efflux of phosphate from both age groups. When cells were preincubated in a phosphate-free medium for 24 hours, the uptake of phosphate was increased to 46% and 24% of their corresponding controls in young and adult cells, respectively. The decreased phosphate uptake and limited adaptation to a phosphate-free medium by the adult renal cells may account for the hypophosphatemia and phosphaturia seen in adult and old animals in vivo.