Molecular cloning, characterization, and expression of a cDNA coding Copper/Zinc superoxide dismutase from black porgy.

A full-length complementary DNA (cDNA) clone encoding a putative copper/zinc superoxide dismutase (Cu/Zn-SOD) was amplified by a Polymerase Chain Reaction (PCR) based technique from cDNA synthesized from black porgy, Acanthopagrus schlegeli, mRNA. Nucleotide sequence analysis of this cDNA clone revealed that it comprised a complete open reading frame coding for 154 amino acid residues. The deduced amino acid sequence showed slightly higher identity (72.8-78.1%) with shark and swordfish Cu/Zn-SOD than with Cu/Zn-SOD from mammalian (68.1-70.7%) and plant (55.5-56.5%) sources. The residues required for coordinating copper and zinc are conserved as they are among all reported Cu/Zn-SOD sequences. The deduced amino acid sequence lacks mitochondria targeting sequence, which suggests that the black porgy cDNA clone encodes a cytosolic Cu/Zn-SOD. The coding region of Cu/Zn-SOD from black porgy was introduced into an expression vector, pET-20b(+), and transformed into Escherichia coli AD494(DE3)pLysS. A predominant achromatic zone was detected by activity staining of native PAGE. This indicates that the Cu/Zn-SOD cDNA clone can express active Cu/Zn-SOD enzyme in E. coli.

Acupuncture and chronic pain management.

INTRODUCTION: Although acupuncture has been widely used to treat a variety of pain conditions, convincing scientific evidence for its efficacy is lacking. Earlier randomised controlled trials attempted to follow a double-blind, placebo-controlled model. This approach has encountered many problems such as the virtual impossibility of blinding the acupuncturist and the uncertainties inherent in choosing control acupuncture points. The objective of this review is to assess if acupuncture is an effective treatment for chronic pain. In addition, a number of key methodological issues that arise in the controlled evaluation of acupuncture will be discussed. METHODS: This review is based on the result of previous reviews, meta-analyses and consensus conference. The search was performed with MEDLINE (from 1966), EMBASE (from 1980) and Cochrane library (1999, volume 1). Only randomised trials of acupuncture (involved needling) for subjects with chronic pain published in English were included. RESULTS: Basic science research has demonstrated convincingly that at least in the context of acute pain, acupuncture's effects are related to the release of a variety of natural opioids. Acupuncture has been shown to be effective for postoperative dental pain. There are reasonable studies showing relief of pain with acupuncture on diverse pain conditions such as menstrual cramps, tennis elbow, low back pain and fibromyalgia. This suggests that acupuncture may have a more general effect on pain. However, there are also studies which provide equivocal results because of design, sample size and other factors. The issue is further complicated by inherent difficulties in the use of appropriate controls, such as placebo and sham acupuncture groups. CONCLUSION: As most of the studies were of poor methodological quality, there is a need for further high quality randomised controlled trials. Future studies should also have larger sample sizes, use a valid acupuncture treatment, and have both a short-term and long-term follow-up.

Acupuncture treatment at Ang Mo Kio Community Hospital--a report on our initial experience.

BACKGROUND: We report our initial experience with acupuncture treatment at the Ang Mo Kio Community Hospital's Acupuncture Research Clinic. PATIENTS: One thousand one hundred and twenty-eight patients received a total of 12,172 acupuncture treatment sessions during the period between September 1995 and December 1996. The majority were Chinese (94%), between 40-70 years (70%) with either dialect or Mandarin (68%) as the main spoken language. However, one third of the patients were English speaking, and educational level did not seem to be a factor among our patients accepting acupuncture treatment. Most had either painful conditions (58%) or stroke-related dysfunction (23%). Amongst the painful conditions, arthritis (25%), low back pain (22%) and other musculo-skeletal pain (12%) were the most common. RESULTS: Prior to treatment, about three-quarter of patients believed acupuncture would benefit them while 40% had tried acupuncture before. After completion of treatment, 70% of all patients considered acupuncture safe, 54% were satisfied with the overall result, 51% felt acupuncture was beneficial, while 54% would recommend acupuncture to others with similar conditions. In a subgroup of patients treated for painful conditions, > 90% reported improvement. Near-syncope occurred in 2 patients (0.18%) or during 2 treatment sessions (0.02%). There was no other acupuncture-related complication. CONCLUSION: Our preliminary experience showed that acupuncture is safe, and appears to be beneficial to patients with painful conditions. An overview of acupuncture treatment is presented, and the issues of safety, efficacy and a need to conduct randomised controlled clinical trials are discussed.

Influence of temperature, pH and naloxone on the antinociceptive activity of Channa striatus (haruan) extracts in mice.

We have previously shown that extracts from the snakehead fish, Channa striatus (haruan), have antinociceptive activity on their own, and enhance morphine antinociception. In furtherance of that study, we investigated the effects of temperature, pH and naloxone on the antinociceptive effects of haruan extracts in the mouse abdominal constriction test. The effects of temperature were only studied on mucus extracts since the fillet extract is prepared at high temperature under pressure. There was a general loss of inhibitory activity from 88.4% at 0 degrees C to 61.1% at 80-100 degrees C, though this was only significant at 80 degrees C. The effects of pH were studied on both extracts. Lowering the mucus extract pH to 6.0 (unadjusted pH 7.4) did not significantly affect its activity, while raising it to 8.0 attenuated its activity. Similarly, pH changes had no effect on the fillet extract except at pH 8.0 which produced apparently lower antinociception. Pretreatment with naloxone (4 mg/kg) had no significant effect on the inhibitory effects of either extract in the abdominal constriction test. These results suggest that the active ingredient(s) of these extracts is (are) relatively stable over a wide range of temperature and between pH 6.0 and 8.0. That would be consistent with the possibility of a glycoprotein, polypeptide or polysaccharide.

Gene transfer via pollen-tube pathway for anti-fusarium wilt in watermelon.

In order to obtain transgenic fusarium wilt resistant watermelon plants, squash DNA was introduced into the ovaries of watermelon plants via the pollen-tube pathway. The introduction of foreign genes into ovaries was accomplished using co-transformation with the CaMV35S-GUS as a marker. Transformed watermelon plants contained integrated copies of the GUS activity and the seeds of transformed progeny produced a blue color when stained with 5-bromo-4-chloro-3-indolyl glucuronide, whereas seeds from untransformed control plants did not. Of 200 transformed seedlings, ten were wilt resistant. The presence of the GUS activity in the genome of stable transgenic seedlings was confirmed by Southern blot analysis. Furthermore, the generation of random amplified polymorphic DNA (RAPD) fingerprints using primers with embedded restriction sites showed amplification products unique to these transgenic plants. Primers OPA-1 and OPA-9 gave distinct band patterns of genomic DNA using the polymerase chain reaction.

Antinociceptive activity of Channa striatus (haruan) extracts in mice.

Haruan, Channa striatus, is a snakehead fish consumed in many parts of the southeast Asian region. It is believed to promote wound healing, as well as reduce post-operative pain. In an attempt to establish the scientific basis for the alleged pain-relieving benefits of this fish, we studied the antinociceptive effects of whole fillet and mucus extracts from haruan in the mouse using the abdominal constriction and tail flick tests. In the abdominal constriction test, the 30 min fillet extract exhibited concentration-dependent inhibition of the writhing response in the 10-50% concentration range, with 20% as the IC50 value. This activity was not dependent on the duration of extraction, with no significant differences among the extracts obtained at durations of 10, 20, 30, 60, 90 and 120 min (range between 45-54% inhibition at 20% concentration). The mucus extract also showed concentration-dependent inhibition of the abdominal constriction response-at the highest concentration used the average inhibition was 68.9%, while IC50 value was 25%. Neither the fillet extract (30 min, 20%) nor the mucus extract (25%) had any demonstrable effect on the tail flick latency on their own, but significantly enhanced the antinociceptive activity of morphine in this assay. Similarly, low concentrations of the mucus and fillet extract enhanced the effects of morphine in the abdominal constriction test. Collectively, these results suggest a scientific basis for the folklore practice of eating haruan fish in the post-operative period for pain relief: Haruan extracts have antinociceptive activity and enhance the activity of other antinociceptive agents.