A case of hypersensitivity to progesterone presenting as an eczematous eruption.

Hypersensitivity to progesterone is a rare condition, and it represents a hypersensitivity reaction to endogenous progesterone. Here we report a case of a woman who presented to our attention for evaluation of a rash for a few years on her posterior elbows, forearms, and right lateral lower extremity. We report this case because it describes a rare clinical entity, with an atypical clinical presentation pemphigoid-like, that is rarely described in literature.

Nephrogenic systemic fibrosis with a spectrum of clinical and histopathological presentation: a disorder of aberrant dermal remodeling.

BACKGROUND: Nephrogenic fibrosing dermopathy (NFD) has emerged as a clinicopathologic entity since 1997 and recently renamed as nephrogenic systemic fibrosis (NSF). The etiology and pathogenesis remain uncertain. Characteristic clinical presentation is described as diffuse thickening and hardening of the skin occurring in patients with renal insufficiency. Typical histological features include proliferation of CD34 positive fibrocytes, increased thick collagen bundles and mucin deposition, without significant inflammatory infiltrate. Variations in clinical presentations have been reported, including papular and plaque-like skin lesions, focal lesion only, as well as systemic involvement. Histological changes can be subtle and non-specific, overlapping with other disease processes and harboring features including calcification and osteoclast-like giant cells with osseous metaplasia. METHODS: We reviewed patients with NSF that presented to our dermatology clinic by chart review, clinical examination and histological examination. Skin biopsy specimens were obtained from all cases. Histopathology evaluations were carried out by three dermatopathologists (AD, BS and GK) independently and the features were compared among all the cases. Special stains and immunohistochemistry study were also performed to highlight the histological features. RESULTS: Seven cases of NSF presented with a spectrum of clinical manifestations, from classic diffuse hardening of the skin to localized linear plaques. On histological examination, proliferation of CD34-positive fibrocytes ranged from sparse to dense, collagen bundles ranged from thin to thick, and the interstitial dermal mucin accumulation ranged from scant-patchy to abundant. In addition, the lesion displayed various degrees of vascular proliferation, inflammatory infiltrates and intensities of CD68 and Factor XIIIa staining. Two cases showed extensive dermal calcification and ossification. CONCLUSION: NSF may present with a spectrum of clinical abnormalities, and exhibit overlapping histopathological features resembling cicatrix and other dermal reparative/regenerative processes. NSF may in fact to be a disorder of aberrant extracellular matrix remodeling in patients with renal insufficiency.

Benzoquinolinediones: activity as insect teratogens.

Morphological abnormalities including extra compound eyes, extra heads, and distally duplicated legs were generated in cricket embryos by treating eggs with single doses of either benz[g]isoquinoline-5,10-dione or benzo[h]quinoline-5,6-dione. Slight structural modifications of the molecules resulted in a loss of teratogenic activity, although embryotoxicity occurred. These potent insect teratogens can be used for analysis of developmental events during embryogenesis.

Preinfusion variables predict the predominant unit in the setting of reduced-intensity double cord blood transplantation.

Double cord blood transplantation (DCBT) may overcome the slow hematopoietic recovery and engraftment failure associated with infusion of a single cord blood unit. In DCBT, only one unit typically contributes to long-term hematopoiesis, but little is known about factors affecting cord predominance. As results from a phase I trial suggested that order of infusion may affect cord predominance, we analyzed the effect of preinfusion variables on chimerism patterns of 38 patients enrolled in the initial study and a subsequent phase II trial. All patients were treated with a reduced-intensity conditioning (RIC) regimen of fludarabine, melphalan and thymoglobulin followed by DCBT. By day 100, 66% of patients had hematopoiesis derived from a single cord blood unit. Higher post-thaw total nucleated cell and CD34+ cell dose were associated with cord predominance and in 68% of patients (P=0.03); the predominant cord blood unit was infused first. Only the post-thaw CD34+ cell dose of the predominant unit predicted time to both neutrophil and platelet engraftment. Although based on a small number of patients, our results identify parameters that may affect cord predominance and engraftment in the setting of DCBT following RIC and suggest possible strategies for selecting infusion order for cord blood units.

Marrow graft rejection by repeated transfusions of allogeneic donor spleen cells.

Many hematological diseases require long-term transfusion support, which causes production of donor-reactive antibodies in sensitized recipients. Sensitized patients are at an increased risk for graft rejection when they undergo allogeneic hematopoietic stem cell transplantation (allo-HSCT). Here, we established a highly sensitized murine model to investigate the mechanism of donor graft rejection. After BALB/c mice were repeatedly transfused with allogeneic spleen cells from C57BL/6 mice, there was a significant increase in complement-dependent cytotoxicity in the serum of sensitized mice. For transplantation, 1 x 10(7) bone marrow cells (BMCs) from C57BL/6 mice were injected into lethally irradiated recipient BALB/c mice. Sensitized mice died between 12 and 15 days post-transplantation, while non-sensitized mice remained alive after 28 days. The hematopoietic recovery rate declined over time in sensitized recipients. The homing trace assay showed a rapid disappearance of donor BMCs in the spleen and bone marrow of sensitized recipients. In addition, the recipient cells and antibodies in the sensitized serum were capable of inducing high level of cell- and complement-mediated cytotoxicity to the donor graft. Our finding may explain the impaired hematopoietic stem cell homing and poor hematopoietic engraftment observed in highly sensitized allo-HSCT patients.

Rationally designed anti-HER2/neu peptide mimetic disables P185HER2/neu tyrosine kinases in vitro and in vivo.

Monoclonal antibodies specific for the p185HER2/neu growth factor receptor represent a significant advance in receptor-based therapy for p185HER2/neu-expressing human cancers. We have used a structure-based approach to develop a small (1.5 kDa) exocyclic anti-HER2/neu peptide mimic (AHNP) functionally similar to an anti-p185HER2/neu monoclonal antibody, 4D5 (Herceptin). The AHNP mimetic specifically binds to p185HER2/neu with high affinity (KD=300 nM). This results in inhibition of proliferation of p185HER2/neu-overexpressing tumor cells, and inhibition of colony formation in vitro and growth of p185HER2/neu-expressing tumors in athymic mice. In addition, the mimetic sensitizes the tumor cells to apoptosis when used in conjunction with ionizing radiation or chemotherapeutic agents. A comparison of the molar quantities of the Herceptin antibody and the AHNP mimetic required for inhibiting cell growth and anchorage-independent growth showed generally similar activities. The structure-based derivation of the AHNP represents a novel strategy for the design of receptor-specific tumor therapies.

Cyclin B1 availability is a rate-limiting component of the radiation-induced G2 delay in HeLa cells.

Irradiation of tumor cells results in a G2 delay, which has been postulated to allow DNA repair and cell survival. The G2 delay after irradiation is marked in HeLa and other cells by delayed expression of cyclin B1. To test whether this depression of cyclin B1 contributes to the G2 delay, we induced cyclin B1 expression in irradiated HeLa cells using a dexamethasone-inducible promoter. Induction of cyclin B1 after radiation abrogated the G2 delay by approximately doubling the rate at which the cells reentered mitosis, whereas dexamethasone itself had no effect. However, overexpression of cyclin B1 did not eliminate the G2 delay in irradiated cells. In unirradiated cells, overexpression of cyclin B1 had no effect on cell cycle progression. Confirmation that reduction of cyclin B1 levels would prolong G2 was provided using antisense oligonucleotides to cyclin B1. These results demonstrate that cyclin B1 levels control the length of the G2 delay following irradiation in HeLa cells but do not exclude additional mechanisms controlling the mitotic delay after irradiation.

The G2 block induced by DNA damage: a caffeine-resistant component independent of Cdc25C, MPM-2 phosphorylation, and H1 kinase activity.

Treatment of cells with agents that cause DNA damage often results in a delay in G2. There is convincing evidence showing that inhibition of p34cdc2 kinase activation is involved in the DNA damage-induced G2 delay. In this study, we have demonstrated the existence of an additional pathway, independent of the p34cdc2 kinase activation pathway, that leads to a G2 arrest in etoposide-treated cells. Both the X-ray-induced and the etoposide-induced G2 arrest were associated with inhibition of the p34cdc2 H1 kinase activation pathway as judged by p34cdc2 H1 kinase activity and phosphorylation of cdc25C. Caffeine treatment restored these activities after either of the treatments. However, the etoposide-treated cells did not resume cycling, revealing the presence of an alternative pathway leading to a G2 arrest. To explore the possibility that this additional pathway involved phosphorylation of the MPM-2 epitope that is shared by a large family of mitotic phosphoproteins, we monitored the phosphorylation status of the MPM-2 epitope after DNA damage and after treatment with caffeine. Phosphorylation of the MPM-2 epitope was depressed in both X-ray and etoposide-treated cells, and the depression was reversed by caffeine in both cases. The results indicate that the pathway affecting MPM-2 epitope phosphorylation is involved in the G2 delay caused by DNA damage. However, it is not part of the caffeine-insensitive pathway leading to a G2 block seen in etoposide-treated cells.

Apoptosis: an early event in metastatic inefficiency.

Whereas large numbers of cells from a primary tumor may gain access to the circulation, few of them will give rise to metastases. The mechanism of elimination of these tumor cells, often termed "metastatic inefficiency," is poorly understood. In this study, we show that apoptosis in the lungs within 1-2 days of introduction of the cells is an important component of metastatic inefficiency. First, we show that death of transformed, metastatic rat embryo cells occurred via apoptosis in the lungs 24-48 h after injection into the circulation. Second, we show that Bcl-2 overexpression in these cells inhibited apoptosis in culture and also conferred resistance to apoptosis in vivo in the lungs 24-48 h after injection. This inhibition of apoptosis led to significantly more macroscopic metastases. Third, comparison between the extent of apoptosis by a poorly metastatic cell line to that by a highly metastatic cell line 24 h after injection in the lungs revealed more apoptosis by the poorly metastatic cell line. These results indicate that apoptosis, which occurs at 24-48 h after hematogenous dissemination in the lungs is an important determinant of metastatic inefficiency. Although prior work has shown an association between apoptosis in culture and metastasis in vivo, this work shows that apoptosis in vivo corresponds to decreased metastasis in vivo.

The farnesyltransferase inhibitor FTI-277 radiosensitizes H-ras-transformed rat embryo fibroblasts.

Many tumor cells have a greater resistance to ionizing radiation than their normal counterparts, suggesting that the development of drugs that can reduce that radioresistance would potentiate the efficacy of radiation therapy. Because activated H-ras expression has been shown to markedly increase radiation resistance in some transformed cells, the inactivation of H-ras would then be predicted to radiosensitize these tumor cells, while leaving normal cells unaffected. H-ras depends for activity upon farnesylation, which can be blocked by farnesylation inhibitors, including the compound FTI-277. In keeping with this prediction, inhibition of H-ras processing using FTI-277 resulted in higher levels of apoptosis after irradiation and increased radiosensitivity in H-ras-transformed rat embryo cells but did not affect control cells. These experiments suggest that farnesylation inhibitors may prove clinically useful as radiosensitizers of tumors that depend on ras function.

Allopurinol ameliorates cardiac function in non-hyperuricaemic patients with chronic heart failure.

OBJECTIVE: This study sought to observe the effects of allopurinol on the cardiac function of non-hyperuricaemic patients with chronic heart failure and determine the safety of allopurinol for clinical applications. PATIENTS AND METHODS: A group of 125 consecutive cases of non-hyperuricaemic patients with chronic heart failure who were treated at Chongqing Emergency Medical Centre between July 2011 and June 2012 were enrolled and were randomly divided into allopurinol (300 mg/day) group (n=62) and control group (n=63). During the six months treatment period, levels of cardiac function, brachial artery endothelial function, inflammatory cytokines, and biochemical markers were routinely examined. RESULTS: After three months of allopurinol treatment, patients exhibited an increase in flow-mediated vasodilatation (FMD) of brachial artery, whereas, after six months of treatment, the cardiac function classification was improved; plasma levels of brain natriuretic peptide and tumour necrosis factor-a were decreased; left ventricular internal diameter was diminished; and the ejection fraction was increased (p<0.01 for all the parameters) in patients. Serum uric acid level was decreased during the treatment period for both groups, with no significant difference between the two groups. Liver and kidney dysfunction was not observed among the study participants, and no significant increase in creatine kinase level was detected for either treatment group. CONCLUSIONS: For non-hyperuricaemic patients with chronic heart failure, the addition of six months of allopurinol therapy was safe and effective. Moreover, in these patients, allopurinol treatment not only can significantly ameliorate the left ventricular function and reduce the level of inflammatory factors but could also improve endothelial function.

Detection of repair activity during the DNA damage-induced G2 delay in human cancer cells.

All eukaryotic cells manifest cell cycle delay after exposure to DNA damaging agents. It has been proposed that such cell cycle checkpoints may allow DNA repair but direct evidence of such activity during the radiation-induced G2 delay has been lacking. We report here that cells arrested in G2 by radiation (2-3 Gy) and etoposide incorporate bromodeoxyuridine (BrdU) at discrete foci in the nucleus. We detected G2 cells with CENP-F, a nuclear protein maximally expressed in G2. Caffeine and okadaic acid, both established radiosensitizers, inhibit the incorporation of BrdU in G2 cells. Radioresistant HT29 and OVCAR cells demonstrate BrdU foci formation more frequently during the G2 delay when compared to the more radiosensitive A2780 cell line. The repair foci formed during G2 may be followed through mitosis and observed in daughter cells in G1. Taken together, these observations are consistent with the detection of DNA repair activity during the radiation-induced G2 delay after relatively low doses of radiation.

Induction of the TRAIL receptor KILLER/DR5 in p53-dependent apoptosis but not growth arrest.

The TRAIL death receptor KILLER/DR5 is induced by DNA damaging agents in wild-type p53-expressing cells. Here we show that, unlike the p53-target CDK-inhibitor p21WAF1/CIP1, the TRAIL death receptor KILLER/DR5 is only induced in cells undergoing p53-dependent apoptosis and not cell cycle arrest. Thus GM glioblastoma cells carrying an inducible MMTV-driven p53 gene undergo cell cycle arrest and upregulate p21 but not KILLER/DR5 expression upon dexamethasone exposure. WI38 normal lung fibroblasts undergoing cell cycle arrest in response to ionizing irradiation also induce p21 but not KILLER/DR5 gene expression. KILLER/DR5 upregulation is also deficient in irradiated lymphoblastoid cells derived from patients with Ataxia Teleangiectasia suggesting a role for the ATM-p53 pathway in regulating KILLER/DR5 expression after DNA damage. Inhibition of transcription by Actinomycin D blocks both KILLER/DR5 and p21 induction in cells undergoing p53-dependent apoptosis. Our results suggest that the p53-dependent transcriptional induction of KILLER/DR5 death receptor is restricted to cells undergoing apoptosis and not cells undergoing exclusively p53-dependent G1 arrest.

Netrin UNC-6 and the regulation of branching and extension of motoneuron axons from the ventral nerve cord of Caenorhabditis elegans.

In the Caenorhabditis elegans embryo, some ventral midline motoneurons extend a process circumferentially to the dorsal midline and a process longitudinally along ventral nerve cord interneurons. Circumferential migrations are guided by netrin UNC-6, which repels motoneuron axons dorsally. Although the motoneuron cell bodies and the longitudinal axons are positioned along UNC-6-expressing interneurons in the ventral nerve cord, the circumferential processes extend only from the motoneuron cell bodies and from defined locations along some longitudinal axons. This implies a mechanism regulates motoneuron branching of UNC-6-responsive processes. We show that expression of unc-6DeltaC, which encodes UNC-6 without domain C, partially rescues circumferential migration defects in unc-6 null animals. This activity depends on the netrin receptors UNC-5 and UNC-40. These results indicate that UNC-6DeltaC can provide the circumferential guidance functions of UNC-6. Furthermore, we show that expression of unc-6DeltaC causes motoneuron branching and the extension of processes from abnormal positions along the ventral nerve cord. This activity is also UNC-5- and UNC-40-dependent. We propose that local interactions mediated by domain C regulate motoneuron branching and responsiveness to the UNC-6 cue.

A new strategy for attachment of antibodies to sterically stabilized liposomes resulting in efficient targeting to cancer cells.

The development of long-circulating formulations of liposomes (S-liposomes), sterically stabilized with lipid derivatives of poly(ethylene glycol) (PEG), has increased the likelihood that these liposomes, coupled to targeting ligands such as antibodies, could be used as drug carriers to deliver therapeutic drugs to specific target cell populations in vivo. We have developed a new methodology for attaching monoclonal antibodies to the terminus of PEG on S-liposomes. A new end-group functionalized PEG-lipid derivative pyridylthiopropionoylamino-PEG- distearoylphosphatidylethanolamine (PDP-PEG-DSPE) was synthesized for this purpose. Incorporation of PDP-PEG-DSPE into S-liposomes followed by mild thiolysis of the PDP groups resulted in formation of reactive thiol groups at the periphery of the lipid vesicles. Efficient attachment of maleimide-derivatized antibodies took place under mild conditions even when the content of the functionalized PEG-lipid in S-liposomes was below 1% of total lipid. The resulting S-immunoliposomes showed efficient drug remote loading, slow drug release rates and increased survival times in circulation compared to liposomes lacking PEG. When antibodies recognizing several different tumor-associated antigens were coupled to the PEG terminus of S-liposomes a significant increase in the in vitro binding of liposomes to the target cells was observed. The binding of S-immunoliposomes containing entrapped doxorubicin to their target cell population resulted in increased cytotoxicity compared to liposomes lacking the targeting antibody.

Attachment of antibodies to sterically stabilized liposomes: evaluation, comparison and optimization of coupling procedures.

Several coupling methods for binding antibodies (Ab) to liposomes have previously been developed. We were interested in examining if some of these methods would be suitable for attaching Ab to long-circulating formulations of liposomes (SL), sterically stabilized with poly(ethylene glycol) (PEG). We studied three 'classical' coupling methods in which Ab was attached at the bilayer surface of SL, and two new coupling methods in which Ab was attached at the PEG terminus. Parameters examined including binding efficiency, antibody surface density, the ability of the immunoliposomes to remote-load the anticancer drug doxorubicin, and the specific binding of the resulting immunoliposomes to target cells. The non-covalent biotin-avidin coupling method resulted in low Ab densities at the cell surface, as did a coupling in method in which maleimide-derivatized Ab was attached to the liposome surface through a thiolated phospholipid incorporated into the liposomes. The low levels of Ab achieved in these method was likely due to interference by PEG with the access of the Ab to the liposome surface. However, when a maleimide-derivatized Ab was coupled to thiolated PEG, moving the coupling reaction away from the liposome surface, very high coupling efficiencies were achieved, and these immunoliposomes achieved good specific binding to their target cells. Oxidizing the Fc region of the Ab and coupling it to the PEG terminus through a hydrazone bond was a less efficient coupling method, but had the advantage of retaining Ab orientation. Efficient remote-loading of doxorubicin was found for immunoliposomes in which Ab was attached at the PEG terminus.

An RME1-independent pathway for sporulation control in Saccharomyces cerevisiae acts through IME1 transcript accumulation.

The RES1-1 mutation was isolated on the basis of its ability to allow MATa/MAT alpha diploid Saccharomyces cerevisiae cells to express a late sporulation-regulated gene, SPR3, in the presence of excess copies of RME1. RME1 is a repressor of meiosis that is normally expressed in cells that lack the a1/alpha 2 repressor encoded by MAT. The RES1-1 mutation also supports sporulation in mat-insufficient diploids. This phenotype does not result from a failure to express RME1 and is not due to activation of the silent copies of mating type information. RES1-1 activates sporulation by allowing IME1 accumulation in all cell types, irrespective of the presence of the MAT products. IME1 is still responsive to RME1 in RES1-1 cells, since double mutants (rme1 RES1-1) that are deficient at MAT can sporulate better than either single mutant. RES1-1 is not an allele of IME1.

Treatment of canine malignancies by 1200 R total body irradiation and autologous marrow grafts.

Twenty-five dogs with malignant lymphoma (L) and 18 dogs with solid, nonhematologic tumors (ST) were treated with 1200 R total body irradiation (TBI). Rescue from the otherwise lethal hemopoietic toxicity by infusion of autologous marrow aspirated before TBI was attempted, and survival, response to TBI, and immune reactivity post-grafting were determined. Eight L dogs survived more than 14 days post TBI and marrow grafting, and 12 out of 19 evaluable dogs showed a decrease of 75 per cent or more in clinically detectable tumor. There was no evident relationship between clinical status or marrow status before TBI and survival of more than 14 days or tumor response to TBI. Seven of the 8 survivors ultimately developed recurrent tumor. Eight ST dogs survived more than 14 days. Only 4 of 14 evaluable ST dogs showed significant clinical response of their tumor to TBI. Humoral and cellular immune reactivity were significantly impaired during the 10-week period following TBI and marrow grafting in all dogs studied. These results indicate that therapy in addition to lethal doses of TBI is necessary to cure spontaneous L or to significantly affect ST in dogs. They also provide baseline data which are necessary to assess the immunotherapeutic effectiveness of allogeneic marrow grafts.