Placental alkaline phosphatase reporter

Of interest are recent observations from Harris laboratory demonstrating that graded neuraminidase concentrations in placental alkaline phosphatase digests yielded up to eight bands (R16). Participation of neuraminidase-sensitive sialic acid in the K -dependent nitrophenyl phosphatase in isolated rat liver plasma membranes (El) has been reported. 

An interesting study on the fate of injected placental alkaline phosphatase (N17) has been reported by Posen et al. (P18), who measured heat-stable alkaline phosphatase. Following injection, there was an initial rapid fall in enzyme activity lasting 3-5 hours and then a less rapid disappearance. After 4 hours, two thirds of the initial activity remained, but it took several weeks for complete disappearance. These kinetics resembled those known for serum albumin labeled with iodine-131. 

This order of evaluation (biochemical, first starch gel, second) is amply justified from our own experiences and the consensus of opinion at a recent symposium on multiple molecular forms of enzymes. In particular, heterogeneity of human intestinal alkaline phosphatase on starch gel was reported by Moss (M34) and Fishman and Kreisher (F9, K25). A certain amount of LPSAP intestinal enzyme may occupy the same position that liver alkaline phosphatase migrates to. On the other hand, more than one band in nonliver positions can be produced by intestine. Another similar situation is seen with placental alkaline phosphatase, which may show three bands. Consequently, the starch-gel data can be correlated with biochemical studies only if the nature and organ source of the preparation are known in advance. 

PXR, in the presence of a CYP3A4 inducer, binds to and activates the responsive elements in a reporter gene construct, leading to higher expression of a reporter gene, which is usually luciferase, chloramphenicol acetyl transferase (CAT), or secretory placental alkaline phosphatase (SPAP). The expression of the reporter gene is determined, and the ratio between the treatment and vehicle control is used to estimate human PXR-dependent induction potential of the test compound. 

Alkaline phosphatase, human placental Whole plant, hydroponic Nicotiana tabacum (tobacco) A. tumefaciens transformation of leaf explant Mannopine synthase (mas2 ) N ot reported 20 Jig day"1 g"1 root dry weight (e) 3 % of total medium protein (e) 72... 

Serum alkaline phosphatase elevations have been reported following administration of salt-poor albumin (B5). Placenta is very rich in a heat-stable alkaline phosphatase, and albumin prepared from placental blood has a high activity of this enzyme. In one cirrhotic patient who received 1-6 units per day of albumin obtained from pooled human blood and/or human placenta, the alkaline phosphatase before infusion was 5 Bodansky units and by the thirteenth day of administration had reached a value of 160 units. The physician administering the albumin at first thought the patient was having a severe toxic liver reaction and stopped the therapy. The alkaline phosphatase then started to go down and within 10 days returned to normal levels. Analysis of the albumin indicated that it contained 470 units of alkaline phosphatase activity and was probably responsible for the observed elevations in the serum enzyme activity. Albumin prepared from venous blood did not cause an alkaline phosphatase elevation, but placenta-albumin caused elevations with a half-life of about 8 days (Ml). 

The nephrotoxicity of 16 continues to generate considerable interest. Orellanine was highly toxic to mice (LD50 = 12.5 mg/kg i.p.)[101] and caused interstitial nephritis and tubular necrosis in mouse kidney [102], A summary of 16-induced changes in renal function and morphology has been reported [103]. In LLC-PKi renal epithelial cell cultures, 16 decreased the activity of alkaline phosphatase and lactate dehydrogenase, and decreased the incorporation of H-leucine and H-thymidine [104]. Orellanine was a noncompetitive inhibitor of renal alkaline phosphatase, but a competitive inhibitor of the intestinal and placental enzymes [105]. In canine kidney MDCK cell cultures, 16, or a metabolite of 16, inhibited protein, RNA and DNA synthesis [106]. 

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