Isolation and characterization of a spontaneously immortalized human breast epithelial cell line, MCF-10.

Two sublines of a breast epithelial cell culture, MCF-10, derived from human fibrocystic mammary tissue exhibit immortality after extended cultivation in low calcium concentrations (0.03-0.06 mM) and floating transfers in low calcium (MCF-10F), or by trypsin-Versene passages in the customary (normal) calcium levels, 1.05 mM (MCF-10A). Both sublines have been maintained as separate entities after 2.3 years (849 days) in vitro and at present have been in culture for longer than 4 years. MCF-10 has the characteristics of normal breast epithelium by the following criteria: (a) lack of tumorigenicity in nude mice; (b) three-dimensional growth in collagen; (c) growth in culture that is controlled by hormones and growth factors; (d) lack of anchorage-independent growth; and (e) dome formation in confluent cultures. Cytogenetic analysis prior to immortalization showed normal diploid cells; although later passages showed minimal rearrangement and near-diploidy, the immortal cells were not karyotypically normal. The emergence of an immortal culture in normal calcium media was not an inherent characteristic of the original tissue from which MCF-10 was derived since reactivated cryo-preserved cells from cultures grown for 0.3 and 1.2 years in low calcium were incapable of sustained growth in normal calcium.

EGF does not enhance corneal epithelial cell motility.

Although it is well known that epidermal growth factor (EGF) accelerates corneal epithelial wound healing by stimulating mitosis, it is also believed that EGF may directly stimulate the motility of individual corneal epithelial cells. We employed three different experimental methods to determine if EGF does indeed enhance the motility of corneal epithelial cells (independent of mitotic effects). First, the effects of EGF on the motility of tissue-cultured rat and rabbit corneal epithelial cells were investigated by a Boyden chamber assay. In rat corneal epithelium, these effects were further investigated by a second method, the agarose drop assay. Both assay techniques demonstrated no increase in corneal epithelial cell motility in the presence of EGF. These findings were corroborated by a third method which consisted of measuring the closure rate of epithelial wounds in organ-cultured rat corneas in the presence and absence of EGF, while concurrently arresting mitosis with colchicine. The wound closure rate before addition of any drug was 0.46 +/- 0.03 mm2/hr. The wound closure rate with EGF (50 ng/ml) was 0.55 +/- 0.03 mm2/hr, significantly (P less than 0.005) more rapid than the drug-free controls. However, when EGF (50 ng/ml) and colchicine (40 micrograms/ml) were used simultaneously, the acceleration of wound closure by EGF was completely negated by the presence of colchicine, resulting in a wound closure rate (0.46 +/- 0.06 mm2/hr) that did not differ significantly (P greater than 0.50) from that of the drug-free control.(ABSTRACT TRUNCATED AT 250 WORDS)

Problems associated with penetrating keratoplasty for corneal edema in congenital glaucoma.

Corneal edema from congenital glaucoma is a rare indication for penetrating keratoplasty. We report the complications and problems associated with eight consecutive penetrating keratoplasties performed in adult eyes with a history of congenital glaucoma. Only 25% of the eyes achieved 20/40 or better vision after surgery. These generally poor results can be attributed to previous glaucomatous optic nerve damage, to complications related to previous surgery, and possibly to amblyopia. The most common surgical complication was postoperative elevation of intraocular pressure, which occurred in all the cases (8/8 eyes), and required permanent augmentation of glaucoma medications in seven eyes (88%), and glaucoma surgery in four eyes (50%). Two eyes (25%) developed corneal graft failure, one from immune rejection and the other from severe postoperative glaucoma necessitating cyclocryotherapy. In view of these complications and the multiple impediments to good postoperative vision, we advise that penetrating keratoplasty be reserved for patients with severe visual disability whose preoperative glaucoma is well controlled.

Lumbar punctures and meningitis.

Four patients, with an additional seven from the literature, had meningitis following a lumbar puncture (LP) that disclosed normal cerebrospinal fluid (CSF). Animal studies demonstrate that perforation of the meninges in the presence of bacteremia enhances the development of meningitis. Simultaneous blood culture should be obtained with all LPs. Regardless of the results of the initial LP, a second CSF examination is recommended in any patient whose clinical condition is deteriorating. If the initial blood culture is positive, a second LP should be strongly considered in all newborn and very young infants.