Analysis of D2 d : a D-region class I gene

The mouse major histocompatibility complex is composed of several genes arranged into the K, D, Qa, and Tla regions. The D region of the BALB/c mouse includes genes D2 ^d , D3 ^d , and D4 ^d , in addition to H-2D ^d and H-2L ^d . We have determined the DNA sequence of the D2 ^d gene and compared it with the known sequences of several class I genes. The exon/intron structure of the D2 ^d gene is similar to other class I genes. It also contains similar 5 regulatory elements. A frameshift occurs in exon seven, resulting in a gene product with a truncated cytoplasmic tail. To examine the surface expression of the D2^d molecule, we generated an exon-shuffled construct containing the promoter and exons 1–3, encoding the signal peptide, 1, and 2 external domains of the D2 ^d gene linked to exons 4–8, encoding the 3, transmembrane and cytoplasmic domains, of the H-2D ^d gene. The construct was transfected into mouse L cells, and a protein was detected at the cell surface by a monoclonal antibody (mAb) specific for the 3 domain of H-2D^d, as well as by other class I-specific mAbs. Although D2^d is expressed at low levels, it may be a functional class I gene that most probably evolved from a Qa region gene.     

Self-harm: 2. Deliberate nonfatal self-harm.

The incidence of deliberate acts of self-harm, such as drug overdoses, has greatly increased in the past two decades. The term "suicide attempt" is really a misnomer. For most people who harm themselves the primary motivation is rarely to die. As well, there are many differences between self-harm patients and patients who have committed suicide. The former tend to be characterologically disturbed and socially disadvantaged. Intervention narrowly focused on suicide prevention will lead to lengthy hospital stays in high-risk cases and ineffective treatment for most self-harm patients, who tend not to comply with outpatient treatment. Crisis-oriented intervention with a brief hospital stay in all cases of self-harm may provide more effective assessment and management.     

Surgical treatment of thyrotoxicosis.

A retrospective study of thyrotoxic patients treated by subtotal thyroidectomy between 2 and 21 years ago in the north-east of Scotland showed that 20% of the patients could not be identified or traced at the time of the survey. The thyroid status of 40% of patients followed up was abnormal.It is now accepted that radioiodine treatment of thyrotoxicosis is followed by a significant incidence of late onset hypothyroidism, and life-long follow-up is regarded as obligatory. The findings in this study indicate that similar methods of aftercare are required for surgically treated patients and for all patients receiving thyroxine-replacement therapy.     

PATHS OF TRANSTUBULAR WATER FLOW IN ISOLATED RENAL COLLECTING TUBULES

The cells of perfused rabbit collecting tubules swell and the intercellular spaces widen during osmotic flow of water from lumen to bath induced by antidiuretic hormone (ADH). Ouabain had no influence on these changes. In the absence of net water flow intercellular width was unaffected when tubules were swollen in hypotonic external media. Therefore, during ADH-induced flow widening of intercellular spaces is not a consequence of osmotic swelling of a closed intercellular compartment containing trapped solutes, but rather is due to flow of solution through the channel. Direct evidence of intercellular flow was obtained. Nonperfused tubules swollen in hypotonic media were reimmersed in isotonic solution with resultant entry of water into intercellular spaces. The widened spaces gradually collapsed completely. Spaces enlarged in this manner could be emptied more rapidly by increasing the transtubular hydrostatic pressure difference. In electron micrographs a path of exit of sufficient width to accommodate the observed rate of fluid flow was seen at the base of the intercellular channel. It is concluded that the intercellular spaces communicate with the external extracellular fluid and that water, having entered the cells across the luminal plasma membrane in response in ADH, leaves the cells by osmosis across both the lateral and basilar surface membranes.     

Sequential protooncogene expression in regenerating kidney following acute renal injury

Following loss of functional renal mass due to acute injury, there is significantly increased proliferation of tubular epithelium to replace injured and necrotic cells. In contrast, following uninephrectomy, the contralateral kidney increases in size primarily by hypertrophy, with little cellular proliferation. We and others have demonstrated only modest increases in renal protooncogene expression following uninephrectomy. In this study, we demonstrate markedly elevated expression of the protooncogenes c-fos, c-myc, c-Ki-ras, and c-Ha-ras following acute renal injury induced by a single large parenteral dose of folic acid. The expression of these genes occurs in a sequential pattern similar to that seen in proliferating cells in culture and in regenerating liver. In addition, we demonstrate elevated levels of histone H4 and beta-actin mRNAs consistent with increased cell proliferation. These data suggest that the molecular mechanisms regulating cell proliferation in the kidney are similar to those in regenerating liver and in cultured cells. In addition, it appears that these events are regulated normally after acute renal injury and following uninephrectomy, since in both instances the levels of protooncogene expression correlate with the degree of cell proliferation. This is in direct contrast to a pathologic renal condition, polycystic kidney disease, in which the level of protooncogene expression is out of proportion to the degree of cell proliferation. Further studies of the molecular correlates of acute renal injury may yield insight into the pathogenesis of this and other clinically important renal disorders.     

Review article. Starch: as simple as A, B, C?

Starch is the main carbohydrate reserve in plants and an important part of our nutrition. Increasingly, it is being seen by industry as a useful raw material to include in foodstuffs and with which to produce other carbon-based polymers. Our understanding of starch biosynthesis and chemistry has advanced rapidly over the last few years, but our knowledge of how this translates into structure and thence into physicochemical properties and function is still lacking. Here, we have reviewed this information with an emphasis on genetics and physical properties, especially using data from the model crop, pea (Pisum sativum L.).     

Immunohistochemical detection of proliferating cells in vivo

Incorporation of the thymidine analogue 5-bromo-2'-deoxyuridine (BrUdR) into newly synthesized DNA provides the basis of a simple technique for identifying proliferating cells. BrUdR was administered to C57BL/6 mice by continuous infusion for 1-7 days, or by intraperitoneal injection for shorter intervals. Various tissue types, including gut, kidney, and liver, were excised, fixed in neutral buffered formalin, and paraffin-embedded for sectioning. De-paraffinized 4-micron tissue sections and bone marrow samples were incubated with an anti-BrUdR antibody and cells that had traversed S-phase during the BrUdR exposure period were identified immunohistochemically. Proliferation and migration of intestinal epithelial cells were identified by antibody staining after continuous in vivo exposure to BrUdR for 1-4 days, and BrUdR incorporation into proliferating marrow cells was detected within 30 min. Tissues such as normal liver, known to have low levels of proliferation, remained unstained after 3 days' exposure to BrUdR. After we established that normal proliferating cells could be identified using this technique, BrUdR was administered to mice bearing B16 melanomas. Again, proliferating tumor cells were clearly identified in histological sections. The nuclei from these paraffin-embedded tumors were also collected for flow cytometric analysis after de-waxing, rehydration, and pepsin treatment. This combination of techniques made possible the comparison in adjacent tissue sections of labeling index, obtained from stained sections, with percentage S-phase, measured using DNA flow cytometry. The % S-phase was consistently higher than the labeling index obtained with immunocytochemistry, and two-parameter DNA vs BrUdR flow cytometry showed that this difference could be accounted for by a population of unlabeled cells with an S-phase DNA content.