A differential mechanical technique for tissue separation, based on the different physical resistance to grinding between mesophyll (M) and bundle sheath (BS) cells, was tested on dicotyledonous C4 plants A triplex canscens, A triplex halimus, Gomphrena globosa, Amaranthus retroflexus, Amaranthus caudatus and Portulaca oleracea. A metal sieve (35 mesh) was placed inside a mortar and pieces of leaves (0.5 cm2) were ground in an aqueous medium on the sieve to obtain a homogenate. The homogenate was at first collected below the sieve and was then filtered through six layers of muslin. Microscopic examination showed that the filtrate was enriched by the M cells and the residue was enriched by BS cells, few of which were broken. The BS cell fraction was then vigorously ground and filtered; this second filtrate was named the BS cell fraction and the first filtrate was named the M cell fraction. Ribulose 1,5-bisphosphate carboxylase (EC 4.1.1.39) (RuBPC) and phosphoenolpyruvate carboxylase (EC 4.1.1.31) (PEPC) were assayed, and chlorophyll determinations and protein estimations were made on both fractions. As expected, PEPC showed higher activities in the M fractions; contrary to expectation RuBPC was present in M cell fractions in the six dicotyledonous C4 plants tested. The relative high RuBPC activities found in the M fraction could not be explained in terms of bundle sheath contamination.
Matrix metalloproteinases (MMPs) as well as their inhibitors (TIMPs) play a crucial role in controlling extracellular matrix turnover and have recently been associated with atherosclerosis, myocardial and vascular injury. Moreover, the genetic variability of MMP genes has been suggested to play an important role in vascular remodeling and age-related arterial stiffening. This study aims to describe associations of 14 selected polymorphisms in genes for MMPs and TIMPs with selected cardiovascular parameters (including central pulse pressure), clinical conditions and drug treatment profiles in 411 stable ischemic patients with preserved systolic function of the left ventricle. The genotyping of 14 single-nucleotide polymorphisms in 8 genes was carried out either using 5′ exonuclease (TaqMan®) reagents or by restriction analysis. Numerous associations of the investigated polymorphisms with systolic and diastolic blood pressure, maximum left ventricular end diastolic pressure and ejection fraction were observed. While some of the observed effects were found to be age-dependent, associations with clinical conditions (hypertension, diabetes mellitus, angina pectoris) were only observed in women and associations with four groups of drugs (statins, nitrates, calcium channel blockers, anti-aggregation drugs) were only observed in men. The results of this study indicate that the genetic variability of MMPs and TIMPs is an important factor which influences cardiovascular functions and may have important consequences for individual therapy customization in the future.