The Role of the Carbohydrate Recognition Domain of Placental Protein 13 (PP13) in Pregnancy Evaluated with Recombinant PP13 and the DelT221 PP13 Variant

Introduction

Placental protein 13 (PP13), a placenta specific protein, is reduced in the first trimester of pregnancy in women who subsequently develop preeclampsia. A naturally occurring PP13 deletion of thymidine at position 221 (DelT₂₂₁ or truncated variant) is associated with increased frequency of severe preeclampsia. In this study we compared the full length (wildtype) PP13 and the truncated variant.

Methods

Full length PP13 or its DelT₂₂₁ variant were cloned, expressed and purified from E-Coli. Both variants were administrated into pregnant rats at day 8 of pregnancy for slow release (>5 days) through osmotic pumps and rat blood pressure was measured. Animals were sacrificed at day 15 or day 21 and their utero-placental vasculature was examined.

Results

The DelT₂₂₁ variant (11 kDA) lacked exon 4 and a part of exon 3, and is short of 2 amino acids involved in the carbohydrate (CRD) binding of the wildtype (18 kDA). Unlike the wildtype PP13, purification of DelT₂₂₁ variant required special refolding. PP13 specific poly- clonal antibodies recognized both PP13 and DelT₂₂₁ but PP13 specific monoclonal antibodies recognized only the wildtype, indicating the loss of major epitopes. Wildtype PP13 mRNA and its respective proteins were both lower in PE patients compared to normal pregnancies. The DelT₂₂₁ mutant was not found in a large Caucasian cohort. Pregnant rats exposed to wildtype or DelT₂₂₁ PP13 variants had significantly lower blood pressure compared to control. The wildtype but not the DelT₂₂₁ mutant caused extensive vein expansion.

Conclusion

This study revealed the importance of PP13 in regulating blood pressure and expanding the utero-placental vasculature in pregnant rats. PP13 mutant lacking amino acids of the PP13 CRD domain fails to cause vein expansion but did reduce blood pressure. The study provides a basis for replenishing patients at risk for preeclampsia by the full length but not the truncated PP13.     

Isolation and characterization of basil mosaic virus in Egypt

Different viral infection symptoms were observed on leaves of basil plants (Ocimum basilicum L.), growing at El-Monofia Governorate, Egypt, during 2015–2016. The majority of these symptoms were mosaic and leaf malformation. The pathogenic agent of this disease is known as basil mosaic virus (BsMV). The sensitive hosts to BsMV are Chenopodium amaranticolor and Phaseolus vulgaris cv. (brown beans). The biological properties of BsMV were found to be 10^?4, 6 days and 66 °C for dilution end point, longevity in vitro and thermal inactivation point, respectively. The electron micrograph of the isolated virus indicated this virus was found to be isometric particles of approximately, 50 nm in diameter and could be classified as tentative member of the Fabavirus. Real-time PCR detection by the Fab primer pairs was confirmed at Ct value = 25.44 for the total cDNA isolated from infected Basil plants and 27.94 for the cDNA of purified virus particles, while CMV primer pairs showed no amplification for both samples. According to the available reports this seems to be the first record for basil mosaic virus in Egypt.     

Improved synthesis and pharmacologic activity of the enantiomers of a new benzofurane type antiarrhythmic compound

An improved synthesis of the enantiomers of the new benzofurane-type antiarrhythmic compound 1 is described, which makes use of the enantiomerically pure mbe-lactol. Thus, acylation of the benzofurane 4 with acetic anhydride and subsequent bromination gave the bromoacetyl-derivative 6 , which, after reduction with LiAlH₄, was protected with mbe-lactol to give a mixture of the diastereomers 8A and 8b . After separation via column chromatography assignment of absolute configuration was carried out using a well-established NMR- method. Reaction with propylamine and cleavage of the protective group gave (R)- 1 and (S)- 1 , respectively. Enantiomeric purity was confirmed using a direct HPLC method with rsp-cyclodextrine as stationary phase. Pharmacological investigations on isolated guinea pig heart muscle preparations showed that GE 68 and its two enantiomers did not significantly differ from each other with regard to their negative inotropic, negative chronotropic, and lack of β-adrenoceptor blocking action. In contrast, the reference drug propafenone was equally potent in its negative inotropic and chronotropic activity as GE 68, but additionally showed a weak β-adrenoceptor blocking activity. © 1994 Wiley-Liss, Inc.     

Depolarization exposes the voltage sensor of the sodium channels to the extracellular region

Summary Two domains of Na channels were mapped with site-specific antibodies raised in rabbit against synthetic peptides corresponding to a part of the voltage sensor of internal repeat 1C ₁ ⁺ (amino acids 210–223) and to a region designated dipole (amino acids 1690–1699) of eel electroplax sodium channels. The antibodies bind to their respective domains in both purified and membrane-bound channels and immunoprecipitate the channels from eel electroplax and rat brain synaptosomes.Anti-C ₁ ⁺ depresses the action potential of rat sciatic nerve in a concentration-dependent way. It binds to the external side of rat brain synaptosomal vesicle, and its binding is potentiated by depolarization. Anti-dipole binds to the inner side of the vesicle, and the binding is inhibited by depolarization.We are most grateful to Dr. M.T. Tosteson (Harvard Medical School) for providing us with samples of the S₄IV peptides. We wish to express our gratitude to Drs. D. Gordon (Hebrew University) and A. Safran (The Weizmann Institute) for helping in the immunoprecipitation procedure, to Drs. H. Rahamimoff (Hebrew University) and A. Barzilai (Columbia University) for advising us with the vesicle experiments, to Drs. D. Kassel and M. Gavish (Technion) for many fruitful discussions, and to Dr. Y. Palti (Technion) for discussions of electric field and suggesting the dipole peptide. This work was supported by a basic research fund (BRF) of The Israel Academy of Sciences #430.87 (H.M. and G.S.), a BSF Grant #84-00367 (H.M.) and The Henry Gutwirt Fund for the Promotion of Research-Technion VPR Fund #184-0093 (H.M.).