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Biomedical subjects

L Luo

Publications and source records attributed to L Luo.

At least 235 records · Page 13Linked to original sources
PubMed1994

Control of expression, glycosylation, and secretion of HIV-1 gp120 by homologous and heterologous signal sequences.

The HIV-1 gp120 signal sequence of 30 amino acids is longer than most glycoprotein signal sequences and contains an average of 5 positively charged amino acids. The HIV-1 gp120 gene with its natural signal sequence expressed in any prokaryotic or eukaryotic expression systems showed extremely low levels of synthesis and secretion. However, deletion of the HIV-1 gp120 signal sequence results in production of large quantities of a nonglycosylated form of gp120 in Spodoptera frugiperda cells. Substitution of the gp120 natural signal sequences with the signal sequences from honeybee mellitin or murine interleukin 3 promotes a high level of expression of a glycosylated form of gp120 and efficient secretion. These heterologous signal sequences contain one (mellitin) or no (IL-3) positively charged amino acids and led us to investigate the role of the positively charged amino acids in the signal sequence of HIV-1 gp120. Four charge-altered forms of the gp120 signal sequence of HIV-1 were constructed by site-directed mutagenesis in which the positively charged amino acids were sequentially substituted with neutral amino acids. The results of these experiments showed that the expression and secretion of gp120 was progressively increased by eliminating the positively charged amino acids in a stepwise fashion. However, the substitution of all positively charged amino acids resulted in the accumulation of nonglycosylated gp120 within the cells with decreased amounts of the glycosylated form of gp120. These results demonstrate that the positively charged amino acids in the signal sequence of HIV-1 gp120 are key factors in determining its poor expression and secretion. Analyses of intracellular transport and folding of gp120 further indicate that the presence of a highly charged, uncleaved signal sequence is an important factor limiting transport of gp120 from the rough ER to the Golgi apparatus.

Amino Acid Sequence↗
PubMed1994

Mapping of functional domains for HIV-2 gag assembly into virus-like particles.

The human immunodeficiency virus type 2 gag precursor protein, pr41, self assembles as virus-like particles (VLP) when the gag gene is expressed in insect cells. To map the functional domains for HIV-2 gag VLP formation, a series of deletion mutants was constructed by removing sequentially the C-terminal region of HIV-2 gag precursor protein and expressing the truncated gag genes in SF9 insect cells by means of recombinant baculoviruses. We found that deletion of up to 143 amino acids at the C-terminus of HIV-2 gag, leaving 376 amino acids at the N-terminus of the protein, did not prevent VLP formation. However, an additional four amino acids deletion from the C-terminus, which represents 372 amino acids at the N-terminus, made gag protein fail to form VLP. There is a proline-rich region at amino acid positions 372 and 377 of HIV-2 gag. To analyze the role of these proline residues, we generated five mutants in which proline was changed sequentially into leucine. Our results showed that replacement of one or two prolines did not stop gag VLP formation, whereas replacement of all three prolines by leucine residues completely abolished VLP assembly. Our data demonstrate that the C-terminal p12 region of HIV-2 gag precursor protein and the zinc finger domain are dispensable for gag VLP assembly, but the presence of at least one of the three proline residues located between amino acid positions 372 and 377 of HIV-2NIH-Z is required.

Animals↗
PubMed1994

The effects of intrathecal morphine and clonidine on the prevention and reversal of spinal cord hyperexcitability following sciatic nerve section in the rat.

We have previously shown that intrathecal (i.t.) morphine, but not the alpha 2-adrenoreceptor agonist clonidine, administered prior to sciatic nerve section, reduced the level of autotomy in rats, which is a behavioural model of neuropathic pain. Neither drug was effective when administered 15 min after nerve section. We now examined the effects of i.t. morphine and clonidine on the development of flexor reflex hyperexcitability following sciatic nerve section in acute physiological experiments. The flexor reflex was recorded from the hamstring muscles in decerebrate, spinalized, unanesthetized rats. The effect of sciatic nerve section on the flexor reflex without drugs was compared with axotomy performed 60 min after i.t. injection of 3 micrograms or 30 micrograms morphine, as well as 50 micrograms clonidine. The effect of these drugs on reversing reflex hyperexcitability was also examined. Both doses of morphine administered prior to sciatic nerve section profoundly depressed the baseline reflex and the higher dose almost completely abolished reflex hyperexcitability following nerve section. In contrast, clonidine pre-administration was less effective in depressing the baseline reflex and blocked reflex hyperexcitability less than morphine. Both morphine and clonidine administered 15 min after nerve section reversed spinal hyperexcitability. Thus, the ability of morphine to prevent the occurrence of autotomy may be related to its effectiveness in blocking axotomy-induced hyperexcitability. These physiological data suggest that even a short period of spinal cord hyperexcitability following nerve injury may lead to the development of neuropathic pain.

Animals↗
PubMed1994

Toward functional magnetic stimulation (FMS) theory and experiment.

This paper examines the use of magnetic fields to functionally stimulate peripheral nerves. All electric fields are induced via a changing magnetic field whose flux is entirely confined within a closed magnetic circuit. Induced electric fields are simulated using a nonlinear boundary element solver. The induced fields are solved using duality theory. The accuracy of these predictions is verified by saline bath experiments. Next, the theory is applied to the stimulation of nerves using small, partially occluded ferrite and laminated vanadium permendur cores. Experiments demonstrate the successful stimulation of peripheral nerves in the African bullfrog with 11 mA, 153 mV excitations. These results offer a new vista of possibilities in the area of functional nerve stimulation. Unlike functional electric stimulation (FES), FMS does not involve any half cell reactions, and thus would not have the commensurate FES restrictions regarding balanced biphasic stimulation, strength duration balances, and oxidation issues, always exercising care that the electrodes remain in the reversible operating regime.

Action Potentials↗
PubMed1994

[Effect of crystal no. 4 of polygonum cuspidatum on the viability of island flaps with venous stasis: an experimental study].

The effects of crystal No. 4 of Polygonum cuspidatum on microcirculatory changes of island flaps in rats following resumption of blood flow after venous occlusion of six hours were observed with a special TV set with magnification of 500X. The pathological events and survival rate of flaps were recorded as well. The results showed that the survival rate, amount of open capillaries, amount of blood flow, and velocity of blood flow of flaps treated by crystal No. 4 of polygonum cuspidatum were much higher than those of untreated control flaps. The white thrombosis and endothelial damage of dermal vessels were alleviated by administration of crystal No. 4 of polygonum cuspsidatum. The experiment also showed that there were no differences between flaps treated with crystal No. 4 of polygonum cuspidatum preoperatively and flaps treated with it postoperatively.

Animals↗
PubMed1993

Low-dose intrathecal clonidine releases tachykinins in rat spinal cord.

In decerebrate, spinalized, unanesthetized rats, 37 pmol (10 ng) intrathecally injected clonidine facilitated the flexor reflex. This effect was blocked by the specific tachykinin antagonists CP-96,345, which acts at the NK1 receptor, and Men 10207, which acts at the NK2 receptor. Thus, low-dose intrathecal clonidine releases the tachykinins substance P and neurokinin A in the spinal cord.

Animals↗
PubMed1993

Acidic and basic FGF mRNA expression in the adult and developing rat cochlea.

In situ hybridization was used to document the distribution of mRNA encoding acidic and basic fibroblast growth factor (aFGF and bFGF) in the rat cochlea from embryonic day (E) 16 to postnatal day (P) > 60. bFGF mRNA was not detected in the cochlea at any age. In the adult, aFGF mRNA was strongly expressed in spiral ganglion (SG) neurons, and this expression increased from base to apex. The stria vascularis (SV) and spiral prominence (SP) showed lesser expression which was equal in all turns. Developmentally, low level expression of aFGF mRNA was first seen in the SG at E-20, and remained low until P-4. Expression increased from P-6 to P-14, when adult levels were reached. aFGF mRNA was also observed in the developing hair cells of all turns at E-20. This expression increased after birth but disappeared after P-6. Expression in the SV and SP was first noted at E-20 and reached adult levels by P-16 and P-10, respectively. High levels of aFGF mRNA in the adult SG suggest that aFGF is important for the maintenance of SG neuron function and structure. aFGF in hair cells during the first postnatal week may be involved in the establishment of cochlear innervation.

Animals↗
PubMed1993

Glycosylation is necessary for the correct folding of human immunodeficiency virus gp120 in CD4 binding.

Conflicting results have been reported regarding the role of carbohydrate on human immunodeficiency virus (HIV) envelope glycoprotein gp120 in CD4 receptor binding. Glycosylated, deglycosylated, and nonglycosylated forms of HIV type 1 (HIV-1) and HIV-2 gp120s were used to examine CD4 receptor-binding activity. Nonglycosylated forms of gp120 generated either by deletion of the signal sequence of HIV-1 gp120 or by synthesis in the presence of tunicamycin failed to bind to CD4. In contrast, highly mannosylated gp120 bound to soluble CD4 molecules well. Enzymatic removal of carbohydrate chains from glycosylated gp120 by endoglycosidase H or an endoglycosidase F/N glycanase mixture had no effect on the ability of gp120 to bind CD4. An experiment which measured the ability of gp120 to bind to CD4 as an assay of the proper conformation of gp120 showed that carbohydrate chains on gp120 are not required for the interaction between gp120 and CD4 but that N-linked glycosylation is essential for generation of the proper conformation of gp120 to provide a CD4-binding site.

CD4 Antigens↗
PubMed1993

Viral liposomes released from insect cells infected with recombinant baculovirus expressing the matrix protein of vesicular stomatitis virus.

The matrix (M) protein of vesicular stomatitis virus (VSV) has been found to promote assembly and budding of virions as well as down-regulating of VSV transcription. Large quantities of M protein can be produced in insect cells infected with recombinant baculovirus expressing the VSV M gene under control of the polyhedrin promoter. Analysis by pulse-chase experiments and density gradient centrifugation revealed that the [35S]methionine-labeled M protein synthesized in insect cells is released into the extracellular medium in association with lipid vesicles (liposomes). Electron microscopy and immunogold labeling showed that M protein expressed in insect cells induced the formation on plasma membrane of vesicles containing M protein, which are released from the cell surface in the form of liposomes. The baculovirus vector itself or recombinants expressing VSV glycoprotein (G) or nucleocapsid (N) protein did not produce the formation of vesicles in infected cells. The baculovirus-expressed M protein retains biological activity as demonstrated by its capacity to inhibit transcription when reconstituted with VSV nucleocapsids in vitro. These data suggest that M protein has the capacity to associate with the plasma membrane of infected cells and, in so doing, causes evagination of the membrane to form a vesicle which is released from the cell. This observation leads to the postulate, which requires further proof, that the VSV M protein can induce the formation and budding of liposomes from the cell membrane surface.

Animals↗
PubMed1992

Chimeric gag-V3 virus-like particles of human immunodeficiency virus induce virus-neutralizing antibodies.

A 41-kDa unprocessed human immunodeficiency virus 2 (HIV-2) gag precursor protein that has a deletion of a portion of the viral protease assembles as virus-like particles by budding through the cytoplasmic membrane of recombinant baculovirus-infected insect cells. We have constructed six different combinations of chimeric genes by coupling the truncated HIV-2 gag gene to the neutralizing domain (V3) or the neutralizing and the CD4 binding domains (V3+CD4BD) of gp120 env gene sequences from HIV-1 or HIV-2. The env gene sequences were inserted either into the middle of the gag gene or at the 3' terminus of the gag gene. Virus-like particles were formed by chimeric gene products only when the env gene sequences were linked to the 3' terminus of the gag gene. Insertion of env gene sequence in the middle of the gag gene resulted in high-level chimeric gene expression but without the formation of virus-like particles. Three different chimeric genes [gag gene with HIV-1 V3 (1V3), gag gene with HIV-2 V3 (2V3), and gag gene with HIV-2 V3+CD4BD (2V3+CD4BD)] formed virus-like particles that were secreted into the cell culture medium. In contrast, the HIV-1 V3+CD4BD/HIV-2 gag construct did not form virus-like particles. The chimeric gag-env particles had spherical morphology and the size was slightly larger than that of the gag particles, but the chimeric particles were similar to the mature HIV particles. Western blot analysis showed that the gag-env chimeric proteins were recognized by antibodies in HIV-positive human serum and rabbit anti-gp120 serum. Rabbit anti-gag 1V3 and anti-gag 2V3 sera reacted with authentic gp120 of HIV-1 and HIV-2, respectively, and neutralized homologous HIV infectivity. Our results show that precursor gag protein has potential as a carrier for the presentation of foreign epitopes in good immunological context. The gag protein is highly immunogenic and has the ability to carry large foreign inserts; as such, it offers an attractive approach for HIV vaccine development.

Animals↗
PubMed1992

A stochastic evolutionary model of molecular sequences.

A stochastic evolutionary model of molecular sequences is proposed. The basic forces in evolution are supposed to be mutation and selection. The concept is somewhat similar to Kauffman-Levin's concept of adaptive walks and corresponding analytical expressions have been developed. The selective force is divided into two parts: a slowly-varying part and a rapidly-changing fluctuation. The latter influences the distribution of sequences and results in an equation of motion along the flow line. The former plays a more important role in the emergence of evolutionary order. It is demonstrated that the asymmetry of selective forces would lead to a definite order of the system.

Animals↗
PubMed1992

Human amyloid precursor protein ameliorates behavioral deficit of flies deleted for Appl gene.

Drosophila amyloid precursor protein-like (Appl) gene encodes a protein product (APPL) similar to beta-amyloid precursor protein (APP) associated with Alzheimer's disease. To understand the in vivo function of APPL protein, we have generated flies deleted for the Appl gene. These flies are viable, fertile, and morphologically normal, yet they exhibit subtle behavioral deficits. We show that a fast phototaxis defect in Appl- flies is partially rescued by transgenes expressing the wild-type, but not a mutant, APPL protein. We further demonstrate a functional homology between APPL and APP, since transgenes expressing human APP show a similar level of rescue as transgenes expressing fly APPL.

Amyloid beta-Protein Precursor↗
PubMed1992

Effects of dietary chloride and magnesium on the incidence of tibial dyschondroplasia in chickens fed on Chinese practical diets.

1. The effects of varying the concentrations of dietary magnesium and chloride on the performance, incidence of tibial dyschondroplasia and tissue element contents of broilers fed on Chinese practical maize soya-bean meal diets was assessed. 2. Two concentrations of dietary chloride (1.5 and 3.5 g/kg) and 3 of dietary magnesium (2.0, 4.0 and 6.0 g/kg) were used to form 6 dietary treatments for a 21-d experimental period. 3. Results from this study indicated that the dietary ingredients of maize and soya-bean meals produced in the northeast of China may predispose or influence the susceptibility of chickens to tibial dyschondroplasia. 4. High dietary chloride induces a high incidence of tibial dyschondroplasia (P less than 0.05) which could be ameliorated by dietary magnesium supplementation. 5. Dietary magnesium plays an anti-rachitic role by improving the deposition of copper and zinc in the bone tissue. 6. Supplementation with magnesium to 4.0 g/kg is recommended for broiler diets in northeast China.

Animal Feed↗
PubMed1991

Expression of firefly luciferase gene in Xenopus laevis oocyte.

Fusion plasmid pSV-Luc20 and pSV-Luc19 were constructed by using the larger BamHI restricted fragments of plasmid pDO432 containing the firefly luciferase (luc) gene and HindIII-BamHI restricted fragment of plasmid pSVK100 containing the SV-40 promoter as insert and vector, respectively. The fusion plasmid had different ligation orientations between the encoding sequence of luc gene and the SV-40 promoter, i.e., they were sense and antisense plasmid. When they were introduced into Xenopus oocyte by micro-injection, only pSV-Luc20 could be transcribed and translated into the enzymatic protein of luciferase with high biological activity. Therefore, firefly luciferase gene and Xenopus oocyte can be used as an excellent system for monitoring the activity of various promoters.

Animals↗
PubMed1991

[The anti-tumor efficacy of natural killer (NK) cells and interleukin 2 against experimental pulmonary metastasis in rats].

Rat spleen cells with NK activity were enriched by discontinuous percoll density gradient centrifugation. Morphologically, about 50% were large granular lymphocytes (LGL), and the majority of these were OX8+ and Asialo GM1+. Cytotoxicity assays in vitro showed that the activity of enriched NK cells was significantly higher than that of unseparated rat spleen cells. Intravenous administration of enriched NK cells plus intraperitoneal crude IL2 led to the successful regression of pulmonary metastasis Walker 256 carcinosarcoma in rats. The effect was much stronger than that caused by unseparated spleen cells plus IL2. The results indicate that adoptive therapy using NK cells instead of spleen cells, in combination with exogenous IL2, might improve the efficiency of tumor immunotherapy.

Animals↗
PubMed1991

[Cerebellar infarction and its assistant examination].

The clinical manifestations of cerebellar infarction are often non-particular and it is difficult to make such a diagnosis clinically. Diagnosis was based on pathology previously. Most of the cases were overlooked while still living and the true morbidity was not known. CT scan has made the diagnosis of this disease possible clinically in some patients, but CT is still not an ideal instrument for diagnosing this disease because of the interference by bony artifacts of posterior fossa. The advent of MRI made it possible to distinguish cerebellar infarction safely and easily. It is not the most powerful tool for diagnosing this disease. The authors reported 15 cases of cerebellar infarction diagnosed by clinical manifestation, neuroimages and pathology and pointed out that further knowledge of this disease is needed.

Adult↗
PubMed1991

The projection linkage between the spinal dorsal horn neurons and both the solitary tract and dorsal column nuclei.

Electrical stimulation of the solitary tract nucleus (SN) and dorsal column nuclei (DCN) as well as microelectrode recording from the lumbal spinal dorsal horn have been used to find and identify the axonal projection of and the afferent innervation on the spinal neurons of pentobarbital-anesthetized rats. A total of 92 neurons was recorded and identified mainly in laminae III-V of the lumbar spinal dorsal horn. Of them, 38 neurons were activated antidromically from stimulation of both the SN and DCN. The other 54 neurons responded synaptically to both the SN and DCN stimulations. The initial antidromic responses of 8 neurons in the first group were followed by one or more responses synaptically driven from the SN and/or DCN stimulation. Conduction velocities were in the range of A delta fibers, but faster in the antidromic responses and slower in the synaptic responses. These results indicate that (i) some spinal neurons issue branched axons of larger-sized A delta fibers and double project to both the SN and DCN; (ii) some of these double projection neurons receive in turn smaller A delta fiber innervation from the SN and/or DCN; and (iii) some other neurons in the spinal cord are dually innervated by smaller A delta fibers originating from both nuclei.

Animals↗