Expression of pro form of prostate-specific antigen by mammalian cells and its conversion to mature, active form by human kallikrein 2

To study the expression, biosynthesis, and processing of prostate-specific antigen (PSA) in mammalian cells, recombinant PSA was expressed in Syrian hamster tumor cell line AV12-664 (AV12-PSA). Expression of PSA was monitored by the Tandem-MP PSA assay. PSA was secreted into the medium during the logarithmic phase of cell growth at >9 microg/ml and was stable. The PSA purified from spent medium of AV12-PSA cells did not exhibit any enzymatic activity and did not complex with the protease inhibitor, alpha-1-antichymotrypsin. These findings indicated that an inactive form of PSA was expressed by AV12-PSA cells. NH2-terminal sequencing confirmed the identity of the PSA purified from the spent medium of AV12-PSA cells to be pro-PSA. This demonstrates that PSA is expressed as pro-PSA by mammalian cells and suggests that pro-PSA may be present in biological fluids. Human kallikrein 2 (hK2), another member of the hK family, is also expressed predominantly in prostate epithelium. Although hK2 has been shown to exhibit trypsin-like activity, little is known about its natural substrates. Using purified proteins, we show that hK2 can convert pro-PSA to mature, enzymatically active PSA, thus establishing a physiological connection between hK2 and PSA. These findings imply that hK2 may be regulating PSA activity in vivo.     

The functional outcome of lower-extremity fractures with vascular injury

Salvage of lower-extremity Gustilo type IIIC fractures is difficult, time-consuming for the patients and physicians, and not universally successful because of poor functional outcomes. Even if successful with limb salvage, the functional result may be unsatisfactory because of mutilating injuries to muscle and nerve, bone loss, and the presence of chronic infection. From July 1991 until July 1994, revascularizations of open IIIC fractures were attempted for wounds with Mangled Extremity Severity Score (MESS) < or = 10. The functional results were evaluated at 2 years after injury. Thirty-six lower-extremity revascularizations were performed on 34 patients, including 1 patient with bilateral distal tibial IIIC fractures and a child with IIIC femoral fracture accompanied by ipsilateral distal tibial amputation. Excluded were patients with below-ankle IIIC fractures as well as patients who underwent immediate amputation at admission. After the revascularization, seven patients with IIIC fractures (7 of 36, 19.4%) underwent secondary amputation within 1 week. At the 2-year follow-up, the overall secondary amputation rate was 25% (9 of 36) and the salvage rate was 75% (27 of 36). Those were no deaths. Of the 29 salvaged limbs among these 27 patients, 23 limbs (23 of 29, 79.3%) required secondary coverage procedures that included 12 free flap transfers (12 of 29, 41.4%). Every patient needed subsequent reconstructive surgery to achieve an acceptable functional result. In this series, MESS was able to predict the secondary amputation rate and the functional result. Sixteen of the 17 limb-salvaged patients with MESS < or = 7 were able to achieve minimal functional requirements, whereas 3 of the 10 patients with MESS = 8 to 10 failed to achieve minimal functional requirements at the 2-year follow-up. Using statistical analysis, we found that the salvaged limbs with MESS < or = 9 exhibited a significant difference in achieving adequate function compared with limbs with MESS > 9. Using our protocol for treatment for IIIC fractures, the threshold for immediate amputation can be raised from MESS = 7 to MESS = 9. Our conclusions are (1) more severely injured limbs have poor functional results, (2) every patient needs subsequent reconstructive surgery, and (3) the MESS may be helpful in decision-making.     

The incidence and characteristics of the clinical picture of recurrent myocardial infarct in the rural population

As many as 3033 patients with myocardial infarction residing in rural areas were kept under observation. In this population, myocardial reinfarction (MRI) was diagnosed in 411 subjects, with 78.8% having had it for 4 years. Those MRI patients ranging between 51 to 60 years showed the greatest prevalence (44.3%). The ratio of micro- to macrofocal (through-and-through) MRI was 1:3, that of males to females 9:1. Microfocal MRI was commonly associated with a pain-free variant of the disease (23.5%) with low frequency of thromboembolic complications (3.1%). Every fifth patient with macrofocal (through-and-through) renecrosis presented with aneurysm of the heart. In a 10-year and longer follow-up, mortality from MRI was 43.6 percent among patients with macrofocal MRI, while deaths attributable to microfocal MRI were estimated to be 28.9 percent. High mortality rates suggest great severity of illness and low efficacy of the drug therapy adopted.     

P2X3 is expressed by DRG neurons that terminate in inner lamina II

The P2X3 receptor subunit, a member of the P2X family of ATP-gated ion channels, is almost exclusively localized in sensory neurons. In the present study, we sought to gain insight into the role of P2X3 and P2X3-containing neurons in sensory transmission, using immunohistochemical approaches. In rat dorsal root ganglia (DRG), P2X3-immunoreactivity (-ir) was observed in small- and medium-sized neurons. Approximately 40% of DRG neuronal profiles in normal rats contained P2X3-ir. In rats that had received neonatal capsaicin treatment, the number of P2X3-positive neurons was decreased by approximately 70%. Analysis of the colocalization of P2X3-ir with cytochemical markers of DRG neurons indicated that approximately 94% of the P2X3-positive neuronal profiles were labelled by isolectin B4 from Bandeiraea simplicifolia, while only 3% contained substance P-ir, and 7% contained somatostatin-ir. In dorsal horn of rat spinal cord, P2X3-ir was observed in the inner portion of lamina II and was reduced subsequent to dorsal rhizotomy, as well as subsequent to neonatal capsaicin treatment. Finally, P2X3-ir accumulated proximal to the site of sciatic nerve ligation, and was seen in nerve fibres in skin and corneal epithelium. In summary, our results suggest that P2X3 is expressed by a functionally heterogeneous population of BSI-B4-binding sensory neurons, and is transported into both central and peripheral processes of these neurons.