Systematic analysis of the MAPK signaling network reveals MAP3K-driven control of cell fate.

The classic network of mitogen-activated protein kinases (MAPKs) is highly interconnected and controls a diverse array of biological processes. In multicellular eukaryotes, the MAPKs ERK, JNK, and p38 control opposing cell behaviors but are often activated simultaneously, raising questions about how input-output specificity is achieved. Here, we use multiplexed MAPK activity biosensors to investigate how cell fate control emerges from the connectivity and dynamics of the MAPK network. Through chemical and genetic perturbation, we systematically explore the outputs and functions of all the MAP3 kinases encoded in the human genome and show that MAP3Ks control cell fate by triggering unique combinations of MAPK activity. We show that these MAPK activity combinations explain the paradoxical dual role of JNK signaling as pro-apoptotic or pro-proliferative kinase. Overall, our integrative analysis indicates that the MAPK network operates as a unit to control cell fate and shifts the focus from MAPKs to MAP3Ks to better understand signaling-mediated control of cell fate.

[Genetic characteristics of hantavirus from rodents in port area of Ningde , Fujian province in the summer of 2020].

Objective: To explore the genetic characteristics and evolution of hantavirus carried by rodents in port area of Ningde in Fujian province in the summer of 2020. Methods: Rodents were captured in the port area of Ningde, the RNA was extracted from rodent lung tissues and detected by using specific kit. The positive samples were used for whole-genome sequencing of the virus. Bioinformatics software was used for the analysis on the similarity and genetic variation of the sequences. Results: A total of 112 rodents were captured, including 5 Rattus norvegicus and 2 Rattus flavipectus, the positive rate of hantavirus was 6.25% (7/112). By virus gene sequencing, two hantavirus complete genome sequences were obtained (named as FJ35 and FJ36, GenBank accession numbers: MW449188-MW449193). The genetic analysis results showed that the hantavirus detected in positive samples were SEOV and shared 99% nucleotide similarity with hantavirus strains LZSF21 and JX20140581 isolated from Shandong province. Phylogenetic analysis using the maximum likelihood method showed that the hantavirus detected in positive samples belonged to S3 subtype, sharing the same subtype with hantavirus strains Z37 from Zhejiang province, LZSF21 from Shandong province, and zy27 and Gongzhuling 415 from northeastern China. Compared with FJ372, the amino acid variation of N259S was observed at sites 251-264 of nucleoprotein, which might be related to antigenicity. Another variation of Q81R was observed in glycoprotein compared with SEOV 80-39 segment of coded amino acid of international reference strain, which might also cause the change in antigenicity. Conclusion: The high positive rate of hantavirus in rodents in the port area of Ningde- would increase the risk of natural human infection and epidemic in local area. The hantavirus positive rodents in this focus might be from an endemic area in Shandong. It is necessary to strengthen the imported rodent control in the port area of Ningde. The virus detected in 2 positive samples belonged to SEOV subtype Ⅲ and shared high homologies of nucleotides and amino acid sequences with the hantavirus strains in surrounding area. However, some slight variations occurred in glycoprotein and nucleoprotein amino acid sequences, which might cause changes in its antigeniity.

Myeloid expression of the AP-1 transcription factor JUNB modulates outcomes of type 1 and type 2 parasitic infections.

Activation of macrophages is a key step in the initiation of immune responses, but the transcriptional mechanisms governing macrophage activation during infection are not fully understood. It was recently shown that the AP-1 family transcription factor JUNB positively regulates macrophage activation in response to Toll-like receptor agonists that promote classical or M1 polarization, as well as to the cytokine interleukin-4 (IL-4), which elicits an alternatively activated or M2 phenotype. However, a role for JUNB in macrophage activation has never been demonstrated in vivo. Here, to dissect the role of JUNB in macrophage activation in a physiological setting, mice lacking JUNB specifically in myeloid cells were tested in two infection models: experimental cerebral malaria, which elicits a pathological type 1 immune response, and helminth infection, in which type 2 responses are protective. Myeloid-restricted deletion of Junb reduced type 1 immune activation, which was associated with reduced cerebral pathology and improved survival during infection with Plasmodium berghei. Myeloid JUNB deficiency also compromised type 2 activation during infection with the hookworm Nippostrongylus brasiliensis, leading to diminished cytokine production and eosinophil recruitment and increased parasite burden. These results demonstrate that JUNB in myeloid cells shapes host responses and outcomes during type 1 and type 2 infections.

Prevalence and risk factors for age-related macular degeneration in the elderly Chinese population in south-western Taiwan: the Puzih eye study.

AIM: This study aimed to ascertain the prevalence of and the risk factors associated with early and late age-related macular degeneration (AMD) among Chinese individuals aged ≥65 years residing in Puzih, Taiwan. METHODS: This population-based cross-sectional study graded digital colour photographs of the ocular fundus of 673 individuals using the Wisconsin Age-Related Maculopathy Grading System. We compared the characteristics of individuals with early and late AMD using χ(2)-analyses and described risk factors for early and late AMD using odds ratios and 95% confidence intervals. RESULTS: Individuals with late AMD were significantly older and more likely to have hypertension. Further, their sunlight exposure time was longer than that of those with early AMD, only drusen, or no AMD lesions (P<0.01). A history of hyperlipidaemia for >10 years was a significant risk factor for early AMD, while old age, hypertension for >10 years, and exposure to sunlight for >8 h per day were associated with late AMD. CONCLUSIONS: The prevalence rate of early AMD in the present study was 15.0%, which is similar to that reported for Caucasians and Japanese included in the European Eye Study and the Hisayama Study, respectively. The late AMD prevalence rate of 7.3% found among our study participants was comparable to that reported by the Greenland Inuit Eye Study and Reykjavik Study, but considerably lower than that reported for Caucasians, indicating that late AMD might be less prevalent among Asians than Caucasians.

Hipk2 cooperates with p53 to suppress γ-ray radiation-induced mouse thymic lymphoma.

A genome-wide screen for genetic alterations in radiation-induced thymic lymphomas generated from p53+/- and p53-/- mice showed frequent loss of heterozygosity (LOH) on chromosome 6. Fine mapping of these LOH regions revealed three non-overlapping regions, one of which was refined to a 0.2 Mb interval that contained only the gene encoding homeobox-interacting protein kinase 2 (Hipk2). More than 30% of radiation-induced tumors from both p53+/- and p53-/- mice showed heterozygous loss of one Hipk2 allele. Mice carrying a single inactive allele of Hipk2 in the germline were susceptible to induction of tumors by γ-radiation, but most tumors retained and expressed the wild-type allele, suggesting that Hipk2 is a haploinsufficient tumor suppressor gene for mouse lymphoma development. Heterozygous loss of both Hipk2 and p53 confers strong sensitization to radiation-induced lymphoma. We conclude that Hipk2 is a haploinsufficient lymphoma suppressor gene.

Amyloid vs FDG-PET in the differential diagnosis of AD and FTLD.

OBJECTIVE: To compare the diagnostic performance of PET with the amyloid ligand Pittsburgh compound B (PiB-PET) to fluorodeoxyglucose (FDG-PET) in discriminating between Alzheimer disease (AD) and frontotemporal lobar degeneration (FTLD). METHODS: Patients meeting clinical criteria for AD (n = 62) and FTLD (n = 45) underwent PiB and FDG-PET. PiB scans were classified as positive or negative by 2 visual raters blinded to clinical diagnosis, and using a quantitative threshold derived from controls (n = 25). FDG scans were visually rated as consistent with AD or FTLD, and quantitatively classified based on the region of lowest metabolism relative to controls. RESULTS: PiB visual reads had a higher sensitivity for AD (89.5% average between raters) than FDG visual reads (77.5%) with similar specificity (PiB 83%, FDG 84%). When scans were classified quantitatively, PiB had higher sensitivity (89% vs 73%) while FDG had higher specificity (83% vs 98%). On receiver operating characteristic analysis, areas under the curve for PiB (0.888) and FDG (0.910) were similar. Interrater agreement was higher for PiB (κ = 0.96) than FDG (κ = 0.72), as was agreement between visual and quantitative classification (PiB κ = 0.88-0.92; FDG κ = 0.64-0.68). In patients with known histopathology, overall classification accuracy (2 visual and 1 quantitative classification per patient) was 97% for PiB (n = 12 patients) and 87% for FDG (n = 10). CONCLUSIONS: PiB and FDG showed similar accuracy in discriminating AD and FTLD. PiB was more sensitive when interpreted qualitatively or quantitatively. FDG was more specific, but only when scans were classified quantitatively. PiB slightly outperformed FDG in patients with known histopathology.

Increased apoptosis, p53 up-regulation, and cerebellar neuronal degeneration in repair-deficient Cockayne syndrome mice.

Cockayne syndrome (CS) is a rare recessive childhood-onset neurodegenerative disease, characterized by a deficiency in the DNA repair pathway of transcription-coupled nucleotide excision repair. Mice with a targeted deletion of the CSB gene (Csb-/-) exhibit a much milder ataxic phenotype than human patients. Csb-/- mice that are also deficient in global genomic repair [Csb-/-/xeroderma pigmentosum C (Xpc)-/-] are more profoundly affected, exhibiting whole-body wasting, ataxia, and neural loss by postnatal day 21. Cerebellar granule cells demonstrated high TUNEL staining indicative of apoptosis. Purkinje cells, identified by the marker calbindin, were severely depleted and, although not TUNEL-positive, displayed strong immunoreactivity for p53, indicating cellular stress. A subset of animals heterozygous for Csb and Xpc deficiencies was more mildly affected, demonstrating ataxia and Purkinje cell loss at 3 months of age. Mouse, Csb-/-, and Xpc-/- embryonic fibroblasts each exhibited increased sensitivity to UV light, which generates bulky DNA damage that is a substrate for excision repair. Whereas Csb-/-/Xpc-/- fibroblasts were more UV-sensitive than either single knockout, double-heterozygote fibroblasts had normal UV sensitivity. Csb-/- mice crossed with a strain defective in base excision repair (Ogg1) demonstrated no enhanced neurodegenerative phenotype. Complete deficiency in nucleotide excision repair therefore renders the brain profoundly sensitive to neurodegeneration in specific cell types of the cerebellum, possibly because of unrepaired endogenous DNA damage that is a substrate for nucleotide but not base excision repair.

Brn3a is a transcriptional regulator of soma size, target field innervation and axon pathfinding of inner ear sensory neurons.

The POU domain transcription factors Brn3a, Brn3b and Brn3c are required for the proper development of sensory ganglia, retinal ganglion cells, and inner ear hair cells, respectively. We have investigated the roles of Brn3a in neuronal differentiation and target innervation in the facial-stato-acoustic ganglion. We show that absence of Brn3a results in a substantial reduction in neuronal size, abnormal neuronal migration and downregulation of gene expression, including that of the neurotrophin receptor TrkC, parvalbumin and Brn3b. Selective loss of TrkC neurons in the spiral ganglion of Brn3a(-/-) cochlea leads to an innervation defect similar to that of TrkC(-/-) mice. Most remarkably, our results uncover a novel role for Brn3a in regulating axon pathfinding and target field innervation by spiral and vestibular ganglion neurons. Loss of Brn3a results in severe retardation in development of the axon projections to the cochlea and the posterior vertical canal as early as E13.5. In addition, efferent axons that use the afferent fibers as a scaffold during pathfinding also show severe misrouting. Interestingly, despite the well-established roles of ephrins and EphB receptors in axon pathfinding, expression of these molecules does not appear to be affected in Brn3a(-/-) mice. Thus, Brn3a must control additional downstream genes that are required for axon pathfinding.

Neurotrophins: roles in neuronal development and function.

Neurotrophins regulate development, maintenance, and function of vertebrate nervous systems. Neurotrophins activate two different classes of receptors, the Trk family of receptor tyrosine kinases and p75NTR, a member of the TNF receptor superfamily. Through these, neurotrophins activate many signaling pathways, including those mediated by ras and members of the cdc-42/ras/rho G protein families, and the MAP kinase, PI-3 kinase, and Jun kinase cascades. During development, limiting amounts of neurotrophins function as survival factors to ensure a match between the number of surviving neurons and the requirement for appropriate target innervation. They also regulate cell fate decisions, axon growth, dendrite pruning, the patterning of innervation and the expression of proteins crucial for normal neuronal function, such as neurotransmitters and ion channels. These proteins also regulate many aspects of neural function. In the mature nervous system, they control synaptic function and synaptic plasticity, while continuing to modulate neuronal survival.

NeuroD-null mice are deaf due to a severe loss of the inner ear sensory neurons during development.

A key factor in the genetically programmed development of the nervous system is the death of massive numbers of neurons. Therefore, genetic mechanisms governing cell survival are of fundamental importance to developmental neuroscience. We report that inner ear sensory neurons are dependent on a basic helix-loop-helix transcription factor called NeuroD for survival during differentiation. Mice lacking NeuroD protein exhibit no auditory evoked potentials, reflecting a profound deafness. DiI fiber staining, immunostaining and cell death assays reveal that the deafness is due to the failure of inner ear sensory neuron survival during development. The affected inner ear sensory neurons fail to express neurotrophin receptors, TrkB and TrkC, suggesting that the ability of NeuroD to support neuronal survival may be directly mediated through regulation of responsiveness to the neurotrophins.

Formation of a full complement of cranial proprioceptors requires multiple neurotrophins.

Inactivation of neurotrophin-3 (NT3) completely blocks the development of limb proprioceptive neurons and their end organs, the muscle spindles. We examined whether cranial proprioceptive neurons of the trigeminal mesencephalic nucleus (TMN) require NT3, brain-derived neurotrophic factor (BDNF) or neurotrophin-4 (NT4) for their development. Complements of TMN neurons and masticatory muscle spindles were decreased by 62% in NT3 null mutants, 33% in BDNF null mutants, and 10% in NT4 null mutant mice at birth. The extent of proprioceptive deficiencies differed among different masticatory muscles, particularly in NT3 null mice. Masticatory muscles of embryonic mice heterozygous for the NT3(lacZneo) or BDNF(lacZ) reporter genes expressed both NT3 and BDNF, consistent with target-derived neurotrophin support of TMN neurons. Although more than 90% of TMN neurons expressed TrkB as well as TrkC receptor proteins by immunocytochemistry in wild-type newborns, TrkC or TrkB null mice exhibited only partial proprioceptive deficiencies similar to those present in NT3 or BDNF;NT4 null mice. Thus, in terms of the survival outcome, two main subpopulations of TMN neurons may exist during embryogenesis, one dependent on TrkC/NT3 functioning and the other utilizing TrkB/BDNF signaling. The differential dependence of TMN neurons on neurotrophins may reflect differential accessibility of the neurons to limiting amounts of NT3, BDNF, or NT4 in target tissues, especially if the tissue distribution or levels of BDNF, NT3, and NT4 were dynamically regulated both spatially and temporally.

POU domain factor Brn-3a controls the differentiation and survival of trigeminal neurons by regulating Trk receptor expression.

Mice lacking the POU domain-containing transcription factor Brn-3a have several neuronal deficits. In the present paper, we show that Brn-3a plays two distinct roles during development of the trigeminal ganglion. In this ganglion, neurons expressing the neurotrophin receptors, TrkB and TrkC, are born between E9.5 and E11.5. In the absence of Brn-3a, very few neurons ever express TrkC, but TrkB-expressing neurons are present at E12.5 in elevated numbers, suggesting that Brn-3a may be a constituent of a regulatory circuit determining which Trk receptor is expressed by these early-born neurons. Most neurons expressing the neurotrophin receptor TrkA are generated between E11.5 and E13.5 in this ganglion and their initial generation is not prevented by absence of Brn-3a. However, after E12. 5, absence of Brn-3a results in a progressive loss in neuronal TrkA and TrkB expression, which leads to a massive wave of apoptosis that peaks at E15.5. Despite complete absence of the Trk receptors at E17. 5 and P0, approximately 30% of the normal complement of neurons survive to birth in Brn-3a mutants. Approximately 70% of these express the GDNF receptor subunit, c-ret; many can be sustained by GDNF, but not by NGF in culture. Thus, the vast majority of surviving neurons are probably sustained in vivo by trophic factor(s) whose receptors are not regulated by Brn-3a. In conclusion, our data indicate the specific functions of Brn-3a in controlling the survival and differentiation of trigeminal neurons by regulating expression of each of the three Trk receptors.

Expression of Trk receptors in the developing mouse trigeminal ganglion: in vivo evidence for NT-3 activation of TrkA and TrkB in addition to TrkC.

Animals lacking neurotrophin-3 (NT-3) are born with deficits in almost all sensory ganglia. Among these, the trigeminal ganglion is missing 70% of the normal number of neurons, a deficit which develops during the major period of neurogenesis between embryonic stages (E) 10.5 and E13.5. In order to identify the mechanisms for this deficit, we used antisera specific for TrkA, TrkB, and TrkC to characterize and compare the expression patterns of each Trk receptor in trigeminal ganglia of wild type and NT-3 mutants between E10.5 and E15.5. Strikingly, TrkA, TrkB, and TrkC proteins appear to be exclusively associated with neurons, not precursors. While some neurons show limited co-expression of Trk receptors at E11.5, by E13. 5 each neuron expresses only one Trk receptor. Neuronal birth dating and cell counts show that in wild-type animals all TrkB- and TrkC-expressing neurons are generated before E11.5, while the majority of TrkA-expressing neurons are generated between E11.5 and E13.5. In mice lacking NT-3, the initial formation of the ganglion, as assessed at E10.5, is similar to that in wild-type animals. At E11.5, however, the number of TrkC-expressing neurons is dramatically reduced and the number of TrkC-immunopositive apoptotic profiles is markedly elevated. By E13.5, TrkC-expressing neurons are virtually eliminated. At E11.5, compared to wild type, the number of TrkB-expressing neurons is also reduced and the number of TrkB immunoreactive apoptotic profiles is increased. TrkA neurons are also reduced in the NT-3 mutants, but the major deficit develops between E12.5 and E13.5 when elevated numbers of TrkA-immunoreactive apoptotic profiles are detected. Normal numbers of TrkA- and TrkB-expressing neurons are seen in a TrkC-deficient mutant. Therefore, our data provide evidence that NT-3 supports the survival of TrkA-, TrkB- and TrkC-expressing neurons in the trigeminal ganglion by activating directly each of these receptors in vivo.

Role of dimerization of the membrane-associated growth factor kit ligand in juxtacrine signaling: the Sl17H mutation affects dimerization and stability-phenotypes in hematopoiesis.

The Kit ligand (KL)/Kit receptor pair functions in hematopoiesis, gametogenesis, and melanogenesis. KL is encoded at the murine steel (Sl) locus and encodes a membrane growth factor which may be proteolytically processed to produce soluble KL. The membrane-associated form of KL is critical in mediating Kit function in vivo. Evidence for a role of cytoplasmic domain sequences of KL comes from the Sl17H mutation, a splice site mutation that replaces the cytoplasmic domain with extraneous amino acids. Using deletion mutants and the Sl17H allele, we have investigated the role of the cytoplasmic domain sequences of KL in biosynthetic processing and cell surface presentation. The normal KL protein products are processed for cell surface expression, where they form dimers. Both Sl17H and the cytoplasmic deletion mutants of KL were processed to the cell surface; however, the rate of transport and protein stability were affected by the mutations. Deletion of cytoplasmic domain sequences of KL did not affect dimerization of KL. In contrast, dimerization of the Sl17H protein was reduced substantially. In addition, we have characterized the hematopoietic cell compartment in Sl17H mutant mice. The Sl17H mutation has only minor effects on hematopoiesis. Tissue and peritoneal mast cell numbers were reduced in mutant mice as well as in myeloid progenitors. Interestingly, long-term bone marrow cultures from Sl17H mice did not sustain the long-term production of hematopoietic cells. In addition, homing of normal hematopoietic progenitors to the spleen of irradiated Sl17H/Sl17H recipient mice was diminished in transplantation experiments, providing evidence for a role of Kit in homing or lodging. These results demonstrate that the membrane forms of KL exist as homodimers on the cell surface and that dimerization may play an important role in KL/Kit-mediated juxtacrine signaling.

Hepatitis B and C coinfections and persistent hepatitis B infections: clinical outcome and liver pathology after transplantation.

Hepatitis B virus (HBV) and hepatitis C virus (HCV) infections are common complications after orthotopic liver transplantation (OLT), but the liver pathology and clinical outcomes of HBV infection with HCV coinfection have not been thoroughly examined. In this study, we used the polymerase chain reaction (PCR) to detect HBV and HCV in pre- and post-OLT sera of 38 patients and correlated the findings with clinical outcome and liver pathology. Of 13 patients who were HBV and HCV negative before OLT, 9 acquired HBV infection, and 4 developed acquired HBV and HCV coinfections after OLT. Persistent HBV infections were present in 10 patients. Three patients with pre-OLT HBV infections developed persistent HBV and acquired HCV coinfections after OLT; 5 with pre-OLT HCV infections developed acquired HBV and persistent HCV coinfections after OLT, and 7 had persistent HBV and HCV coinfections before and after OLT. Portal/periportal inflammation was the same in all groups; however, lobular inflammation and fibrosis were more severe in patients with persistent HBV infections and in those with acquired HBV and HCV coinfections. Two major histopathological patterns were present in patients with HBV and HCV coinfections, one with predominant features of HCV infection, and the other with those of HBV infection. Patients with post-OLT HBV and HCV coinfections had survival rates similar to those with acquired HBV infection, whereas patients with persistent HBV infections experienced more allograft loss caused by chronic hepatitis or fibrosing cytolytic hepatitis, and had a more dire clinical outcome than the others. Although the limited numbers reported in this study prevent a definitive conclusion, our data suggest that in patients with HBV and HCV coinfections, the presence of HCV may improve the clinical outcome as compared with the expected outcome of persistent HBV infection alone.

The murine steel panda mutation affects kit ligand expression and growth of early ovarian follicles.

Mutations at the murine steel (Sl) locus encoding the ligand for the c-kit receptor result in defects in gametogenesis, hematopoiesis, and melanogenesis. Steel Panda (Slpan) is an allele at the Sl locus obtained by an X-ray mutagenesis protocol. Slpan/Slpan homozygotes are mildly anemic black-eyed whites with pigmented ears and scrotum; females are sterile while males are fertile. To investigate the basis of the phenotype of the Slpan mutation, the coding region of the kit ligand (KL) in Slpan/Slpan animals was characterized and shown to be identical to that from +/+ mice. RNA expression patterns in adult Slpan/Slpan mice were investigated by RNA blot analysis and RNase protection assays. KL RNA expression was shown to be reduced in several tissues including testis, lung, and kidney, to about 60% in heterozygotes and 20% in homozygous mutant mice. Intermediate effects were seen in cerebellum and spleen, while in heart and brain no change was apparent. Therefore, the Slpan mutation affects KL RNA levels in a tissue-specific manner. Histological analysis showed that the number of oocytes in neonatal homozygotes was reduced to 20% of that in heterozygotes, and that in juvenile and adult mice ovarian follicle development was arrested at the one-layered cuboidal stage, with a few exceptions. KL production by central cords of the perinatal ovary was severely reduced as shown by immunohistochemistry. In neonatal testes of homozygotes, the germ cell number was reduced to 30% of that in heterozygotes, but meiotic spermatocytes were produced on schedule in juvenile animals. Therefore, a reduced level of KL in Slpan/Slpan ovary arrests ovarian follicle development, while a similar reduction in testes has relatively little effect on spermatogonial development.

The expression pattern of the c-kit ligand in gonads of mice supports a role for the c-kit receptor in oocyte growth and in proliferation of spermatogonia.

The tyrosine kinase receptor c-kit and its ligand KL are required for postnatal development of germ cells, in addition to their role in primordial germ cells. To clarify their function, a detailed examination of the pattern of expression of KL in postnatal gonads was undertaken. In ovaries, the expression of KL as seen by RNA blot analysis and by RNase protection assays is relatively high at birth (P0), low from P5 to P8, and high from P12 onward. KL expression is relatively high in testes of all ages. The forms of KL RNA present in the testes suggest that from P5 onward the membrane-bound form of KL predominates, while in the ovary significant amounts of both forms are present. As observed by in situ hybridization and immunohistochemistry, in the newborn ovary KL is highly expressed in central cords whose cells contribute to the formation of central growing follicles. Expression is low in follicle cells of small growing follicles and increases to high levels in three-layered follicles during late oocyte growth. Large amounts of the ligand are found within growing oocytes. After oocyte growth ceases, expression continues only in the outer layers of multilayered follicles. In the testis, from P0 through P9, KL expression is distinct in Sertoli cells, but not in germ cells. Thereafter, the intensity of KL expression declines as the number of spermatogenic cells increases within the tubules. KL in Sertoli cells appears to be concentrated basally at the stage of the cycle of the seminiferous epithelium when it is known to interact with differentiating type A spermatogonia. These data are consistent with a role for KL in oocyte growth and in facilitating proliferation and/or differentiation of type A spermatogonia.

Early and long-term effects of colectomy and endorectal pullthrough on bile acid profile.

OBJECTIVE: Although total colectomy with mucosal proctectomy and endorectal pullthrough affects two sites critical to the enterohepatic circulation of bile acids, little information is available regarding the manner in which normal digestive physiology is altered by these procedures. This study defines the early and long-term effects of colectomy and endorectal pullthrough on bile acid profile and the long-term effects on biliary lipid metabolism. SUMMARY BACKGROUND DATA: Specific changes in bile acid absorption have been reported in patients after ileal resection. Recent studies from our laboratory indicate that in the early postoperative period, colectomy with endorectal pullthrough causes a significant decrease in gallbladder bile concentrations of total bile acids, cholesterol, phospholipids, and calcium. The observation by several authors that the pouch undergoes morphologic and perhaps functional adaptation suggest that these changes may be transient and perhaps reversible. METHODS: These studies were done in an awake, unanesthetized canine model that allows periodic sampling of gallbladder bile without creation of an external biliary fistula and its associated sequelae. Animals were ultimately randomly assigned to either laparotomy and gallbladder cannulation (N = 6), or gallbladder cannulation with total colectomy and ileorectal anastomosis (N = 7), or biliary cannulation, colectomy, mucosal proctectomy and endorectal pullthrough with ileal reservoir (N = 5). RESULTS: Six weeks after operation, colectomy and ileorectal anastomosis were associated with a significant alteration in the relative composition of bile acids in gallbladder bile. These early changes were manifested by a significant (p < 0.05) increase in taurocholic acid and a concomitant decrease in taurodeoxycholic acid. These changes became even more pronounced in the ileorectal anastomosis group 12 weeks after colectomy and ileorectostomy. Although similar changes in the relative concentrations of individual bile acids occurred in the 6-week endorectal animals, bile acid profile was restored to normal by 12 weeks. CONCLUSIONS: Colectomy with ileorectal anastomosis leads to early and significant changes in bile acid profile, which persist and become even more pronounced with time. In contrast, the construction of an ileal reservoir after colectomy facilitates restoration of a normal bile acid profile. We propose that these alterations in bile acid metabolism result from adaptation of the ileal reservoir as its mucosa assumes functional characteristics of normal colon.

The kit-ligand (steel factor) and its receptor c-kit/W: pleiotropic roles in gametogenesis and melanogenesis.

The c-kit receptor tyrosine kinase belongs to the PDGF/CSF-1/c-kit receptor subfamily. The kit-ligand, KL, also called steel factor, is synthesized from two alternatively spliced mRNAs as transmembrane proteins that can either be proteolytically cleaved to produce soluble forms of KL or can function as cell-associated molecules. The c-kit receptor kinase and KL are encoded at the white spotting (W) and steel (Sl) loci of the mouse, respectively. Mutations at both the W and the Sl locus cause deficiencies in gametogenesis, melanogenesis and hematopoiesis. The c-kit receptor is expressed in the cellular targets of W and Sl mutations, while KL is expressed in their microenvironment. In melanogenesis, c-kit is expressed in melanoblasts from the time they leave the neural crest and expression continues during embryonic development and in the melanocytes of postnatal animals. In gametogenesis c-kit is expressed in primordial germ cells, in spermatogonia, and in primordial and growing oocytes, implying a role at three distinct stages of gametogenesis. Many mutant alleles are known at W and Sl loci and their phenotypes vary in the degree of severity in the different cellular targets of the mutations. While many W and Sl alleles severely affect primordial germ cells (PGC), several mild Sl alleles have weak effects on PGCs and exhibit differential male or female sterility. Steel Panda (Sl(pan)) is a KL expression mutation in which KL RNA transcript levels are reduced in most tissues analyzed. In female Sl(pan)/Sl(pan) mice, ovarian follicle development is arrested at the one layered cuboidal stage as a result of reduced KL expression in follicle cells, indicating a role for c-kit in oocyte growth. Wsh is a c-kit expression mutation, which affects mast cells and melanogenesis. While the mast cell defect results from lack of c-kit expression, the pigmentation deficiency appears to stem from ectopic c-kit receptor expression in the somitic dermatome at the time of migration of melanoblasts from the neural crest to the periphery. It is proposed that the ectopic c-kit expression in Wsh mice affects early melanogenesis in a dominant fashion. The "sash" or white belt of Wsh/+ animals and some other mutant mice is explained by the varying density of melanoblasts along the body axis of wild-type embryos.