Expression of Cre recombinase in chondrocytes causes abnormal craniofacial and skeletal development

The cranial base synchondroses are growth centers that drive cranial and upper facial growth. The intersphenoid synchondrosis (ISS) and the spheno-occipital synchondrosis (SOS) are two major synchondroses located in the middle of the cranial base and are maintained at early developmental stages to sustain cranial base elongation. In this study, we report unexpected premature ossification of ISS and SOS when Cre recombinase is activated in a chondrocyte-specific manner. We used a Cre transgenic line expressing Aggrecan enhancer-driven, Tetracycline-inducible Cre (ATC), of which expression is controlled by a Col2a1 promoter. Neonatal doxycycline injection or doxycycline diet fed to breeders was used to activate Cre recombinase. The premature ossification of ISS and/or SOS led to a reduction in cranial base length and subsequently a dome-shaped skull. Furthermore, the mice carrying either heterozygous or homozygous conditional deletion of Tsc1 or Fip200 using ATC mice developed similar craniofacial abnormalities, indicating that Cre activity itself but not conditional deletion of Tsc1 or Fip200 gene, is the major contributor of this phenotype. In contrast, the Col2a1-Cre mice carrying Cre expression in both perichondrium and chondrocytes and the mice carrying the conditional deletion of Tsc1 or Fip200 using Col2a1-Cre did not manifest the same skull abnormalities. In addition to the defective craniofacial bone development, our data also showed that the Cre activation in chondrocytes significantly compromised bone acquisition in femur. Our data calls for the consideration of the potential in vivo adverse effects caused by Cre expression in chondrocytes and reinforcement of the importance of including Cre-containing controls to facilitate accurate phenotype interpretation in transgenic research.

     

Nitrification using polyvinyl alcohol-immobilized nitrifying biofilm on an O2-enriching membrane

A combination of cell immobilization and membrane aeration approaches was used in a biological reactor to treat NH₄ ⁺ in wastewater. Nitrifying microorganisms, immobilized by polyvinyl-alcohol (PVA) and attached to the surface of a silicone membrane tube, were used to develop a novel reactor for nitrification. The immobilized biofilm had a rubber-like elasticity and resisted shear stress over 5 months of operation. The reactor removed 95% of ammonium, added at 1.97 g N m^–2 d^–1, with O_–2 enriching the membrane.     

Nitrogen removal from wastewater using a double-biofilm reactor with a continuous-flow method

Wastewater microorganisms of nitrification and denitrification were cultivated to compose two biofilm modules, termed the permeable support bioreactor (PSB) and the membrane feeding substrate bioreactor (MFSB). PSB and MFSB were combined in a single tank to develop a double-biofilm reactor, which was used to treat nitrogen contaminants in wastewater. With a membrane supplement of substrates (O(2) and CH(3)OH), the D.O. and COD levels were at a low value in the bulk solution thus inhibitive effects between nitrification and denitrification were minimized. Simultaneous nitrification/denitrification was conducted in the reactor and the double-biofilm reactor achieved high nitrification and denitrification efficiency, of 96.5% and 82%, respectively.